（共享）免疫组织化学染色FAQ

Staining, histochemistry and histotechnology

compiled by

J. A. Kiernan,
Department of Anatomy and Cell Biology,
The University of Western Ontario,
London, Canada

<center> <font><strong>Table of Contents</strong></font></center>


FIXATION, FREEZING ETC

~undefined_Carbodiimides_as_fixatives~Kbr_~H~M~2~undefined_Carnoy_~8amp~J_alcoholic_fixatives~Kbr_~H~M~2~undefined_Perfusion_fixative_for_electron_microscopy~Kbr_~H~M~2~undefined_Fixation_of_frozen_sections.~Kbr_~H~M~2~undefined_Non~Fformaldehyde_commercial_fixatives~Kbr_~H~M~2~undefined_Glutaraldehyde_and_immunohistochemistry~Kbr_~H~M~2~undefined_Isopentane~I_alternative_names~Kbr_~H~M~2~undefined_Lidocaine_in_perfusion_fixation~Kbr_~H~M~2~undefined_Michel~9s_fluid_for_transporting_cells_or_specimens~Kbr_~H~M~2~undefined_Microwave_ovens~I_Advice_for_new_users~Kbr_~H~M~2~undefined_Paraformaldehyde~I_why_won~9t_it_dissolve~N~Kbr_~H~M~2~undefined_Saccomano~9s_fixative~Kbr_~H~M~2~undefined_Zinc~Fcontaining_fixatives~I_What_has_been_published~N~Kbr_~H~M~2~undefined_Alternatives_to_mercury~Fcontaining_fixatives~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1PROCESSING~E_DECALCIFYING~E_EMBEDDING~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Solvent_to_replace_xylene_AND_alcohols~Kbr_~H~M~2~undefined_2~Fbutoxyethanol_~A~4Clereum~4~B_dehydrating_or_clearing_agent~Kbr_~H~M~2~undefined_Decalcification~I_Acid_or_EDTA~N~Kbr_~H~M~2~undefined_Testing_for_completeness_of_decalcification~Kbr_~H~M~2~undefined_Fatty_specimens~I_Processing_into_paraffin.~Kbr_~H~M~2~undefined_Polymethyl_methacrylate_embedding_for_bone~Kbr_~H~M~2~undefined_Mold_release_spray~Kbr_~H~M~2~undefined_Paraffin_processing_of_skin~Kbr_~H~M~2~undefined_Cryoprotection_of_specimens~Kbr_~H~M~2~undefined_Cutting_sections_of_toe_or_finger_nails~Kbr_~H~M~2~undefined_Paraffin_wax~I_crystals~E_additives_and_cutting~Kbr_~H~M~2~undefined_Xylene_substitutes~I_what_are_they~N~Kbr_~H~M~2~undefined_Test_for_water_in_used_absolute_alcohol~Kbr_~H~M~2~undefined_Molecular_sieves_for_making_anhydrous_solvent~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1SECTIONING~E_SLIDE_ADHESIVES~E_MOUNTING~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Sections_coming_off_slides._Which_adhesive~N~Kbr_~H~M~2~undefined_Apathy~9s_mounting_medium_and_variants~Kbr_~H~M~2~undefined_Silanized_~AAPES_or_TES_or_positively_charged~B_slides~Kbr_~H~M~2~undefined_Polishing_undecalcified_bone_sections.~Kbr_~H~M~2~undefined_Polylysine~Fcoated_slides~Kbr_~H~M~2~undefined_Wrinkles_in_plastic_sections~Kbr_~H~M~2~undefined_Wrinkles_in_paraffin_sections_containing_cartilage~Kbr_~H~M~2~undefined_Thick_paraffin_sections~Kbr_~H~M~2~undefined_Sectioning_plastic~Fembedded_specimens~Kbr_~H~M~2~undefined_Iodine_for_removing_mercury_deposits~Kbr_~H~M~2~undefined_Labeling_slides~Kbr_~H~M~2~undefined_Sectioning_plant_material~I_some_hints.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1STAINING_METHODS~E_HISTOCHEMISTRY~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Making_aldehyde~Ffuchsine~Kbr_~H~M~2~undefined_Phosphatases_in_decalcified~E_embedded_tissue.~Kbr_~H~M~2~undefined_Congo_red_for_amyloid~Kbr_~H~M~2~undefined_Cartilage_staining_with_safranine~Kbr_~H~M~2~undefined_Stain_for_Chlamydia_~ACastaneda~9s_method~B.~Kbr_~H~M~2~undefined_Which_staining_method_for_copper_is_best~N~Kbr_~H~M~2~undefined_Diastase_~Aamylase~B_control_for_glycogen~Kbr_~H~M~2~undefined_Evans_blue~E_trypan_blue_and_eosin_as_tracers.~Kbr_~H~M~2~undefined_Gallyas~9_stain~Kbr_~H~M~2~undefined_Gram_staining_of_sections_~ABrown_~8amp~J_Hopps_method~B.~Kbr_~H~M~2~undefined_Oxidants_for_hematoxylin~Kbr_~H~M~2~undefined_McFaydean~9s_stain_for_anthrax_bacilli~Kbr_~H~M~2~undefined_Microglia_with_Griffonia_lectin.~Kbr_~H~M~2~undefined_Picro~Fsirius_red_staining~Kbr_~H~M~2~undefined_Iron_hematoxylin~I_ripening_not_needed.~Kbr_~H~M~2~undefined_Enzyme_histochemistry_on_cell_cultures~Kbr_~H~M~2~undefined_Malachite_green_in_stain_for_Cryptosporidium~Kbr_~H~M~2~undefined_Confusing_dye_names_~Alissamine_fast_red_as_an_example~B~Kbr_~H~M~2~undefined_Mayer~9s_and_Gill~9s_hematoxylins~Kbr_~H~M~2~undefined_Effects_of_pH_on_staining_by_dyes~Kbr_~H~M~2~undefined_Histochemical_stain_for_arsenic~Kbr_~H~M~2~undefined_Giemsa_staining_of_blood_smears~I_several_hints~Kbr_~H~M~2~undefined_Automated_H_~8amp~J_E_staining_problems~Kbr_~H~M~2~undefined_Verhoeff~9s_stain_for_myelin_and_elastin~Kbr_~H~M~2~undefined_Acridine_orange_method_for_DNA_and_RNA~Kbr_~H~M~2~undefined_Quickly_finding_something_in_a_newly_cut_section~Kbr_~H~M~2~undefined_Fluorescent_lectins~I_general_method~Kbr_~H~M~2~undefined_Methyl_blue_and_methylene_blue~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1IMMUNOHISTOCHEMISTRY~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Paraffin_or_frozen_sections_for_immunohistochemistry~Kbr_~H~M~2~undefined_Inhibiting_endogenous_peroxidase~Kbr_~H~M~2~undefined_Using_mouse_primary_antibodies_on_mouse_tissues~Kbr_~H~M~2~undefined_Antigen_retrieval~I_A_patented_or_copyright_phrase~N~Kbr_~H~M~2~undefined_p53_protein~Kbr_~H~M~2~undefined_Prevention_of_fluorescence_fading~Kbr_~H~M~2~undefined_Background_in_immunostained_cartilage~Kbr_~H~M~2~undefined_Endogenous_biotin_in_mast_cells~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1MISCELLANEOUS_STUFF~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Disposal_of_used_diaminobenzidine_~ADAB~B_solutions~Kbr_~H~M~2~undefined_Dilution_of_concentrated_acids~I_formula_etc.~Kbr_~H~M~2~undefined_Disposal_of_waste_from_~4special_stains~4~Kbr_~H~M~2~undefined_Magnification_of_a_photomicrograph~Kbr_~H~M~2~undefined_Can_a_method_be_both_published_and_patented~N~Kbr_~H~M~2~undefined_Books_and_articles_about_artifacts_in_histology~Kbr_~H~M~2~undefined_How_dangerous_is_picric_acid~N~Kbr_~H~M~2~undefined_Which_color_print_film_for_photomicrography~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1FIXATION~E_FREEZING_ETC~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Carbodiimides_as_fixatives~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Does_anyone_know_what_carbodiimide_is_and_how_it_works~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1The_name_~4carbodiimide~4_is_sometimes_applied_to_cyanamide~Kbr_~H~M~2~1~Ahydrogen_cyanamide~E_H2NCN._Don~9t_confuse_this_with_calcium~Kbr_~H~M~2~1cyanamide~E_CaNCN.~B~E_which_does_not_seem_to_have_been_used_as_a~Kbr_~H~M~2~1fixative.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Carbodiimides_are_compounds_that_combine_with_and_cross~Flink~Kbr_~H~M~2~1carboxyl_groups._They_fix_proteins_by_joining_together_C~Ftermini~Kbr_~H~M~2~1and~Hor_side_chains_of_glutamic_and_aspartic_acid_units._Their~Kbr_~H~M~2~1general_chemical_formula_is_R~FN~LC~LN~FR~9~Kbr_~H~M~2~1~Kbr_~H~M~2~1In_contrast~E_aldehydes_combine_mainly_with_protein_nitrogen~Kbr_~H~M~2~1atoms._Cross~Flinks_between_the_lysine_side~Fchain_amino_group~Kbr_~H~M~2~1and_the_amide_nitrogens_of_peptide_linkages_are_thought_to~Kbr_~H~M~2~1do_most_of_the_fixing.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Various_carbodiimides_have_been_used_as_fixatives_over_the~Kbr_~H~M~2~1years~E_but_they_have_never_caught_on_in_a_big_way._They_are~Kbr_~H~M~2~1the_sort_of_things_used_when_more_ordinary_compounds_are~Kbr_~H~M~2~1unsuitable._See_Pearse~9s_Histochemistry~E_Vol._1_~A3rd_ed.~E~Kbr_~H~M~2~1Churchill~FLivingstone~E_Edinburgh~E_1980~B_page_107_for_a~Kbr_~H~M~2~1proper_account.~Kbr_~H~M~2~1~Kbr_~H~M~2~1If_the_antigenicity_of_a_protein_is_critically_dependent_on~Kbr_~H~M~2~1free_amino_groups_of_an_epitope~E_then_one_of_the_carbodiimide~Kbr_~H~M~2~1fixatives_might_be_a_sensible_alternative_to_formaldehyde._If~Kbr_~H~M~2~1it~9s_for_paraffin_sections~E_a_chemically_unreactive_fixative~Kbr_~H~M~2~1such_as_Clarke~9s_or_Carnoy~9s_might_be_even_more_sensible.~Kbr_~H~M~2~1~Kbr_~H~M~2~1John_A._Kiernan~E~Kbr_~H~M~2~1Department_of_Anatomy_~8amp~J_Cell_Biology~E~Kbr_~H~M~2~1The_University_of_Western_Ontario~E~Kbr_~H~M~2~1LONDON~E_Canada_N6A_5C1~Kbr_~H~M~2~1~Kbr_~H~M~2~1Back_to_Table_of_Contents~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Carnoy_~8amp~J_alcoholic_fixatives~Kbr_~H~M~2~1~Kbr_~H~M~2~1Note~I_The_answer_to_Question_2_discusses_the_suitability_of~Kbr_~H~M~2~1alcoholic_and_other_fixatives_for_immunohistochemistry.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question_1.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Any_thoughts_on_the_shelf_life~Hkeeping~Kbr_~H~M~2~1qualities_of_Carnoy~9s_fixative~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1I_always_make_Carnoy~9s_fixative_fresh_just_before_use.~Kbr_~H~M~2~1Otherwise_you_will_find_that_the_fixing_properties_will_vary_if~Kbr_~H~M~2~1the_solution_is_kept_for_any_length_of_time._Making_up_a_fresh~Kbr_~H~M~2~1solution_really_only_takes_a_few_minutes_unless_you_are_talking~Kbr_~H~M~2~1about_Lebrun~9s_modification_in_which_the_solution_is_saturated~Kbr_~H~M~2~1with_mercuric_chloride.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Barry_Rittman~Kbr_~H~M~2~1brittman~PAT~Qmail.db.uth.tmc.edu~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question_2~Kbr_~H~M~2~1~Kbr_~H~M~2~1Are_alcoholic_fixatives_suitable_for_immunohistochemistry~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Fixatives_containing_ethanol_are_generally_not_all_that_great~Kbr_~H~M~2~1for_IHC._About_4~F5_years_ago_we_experimented_with_several~Kbr_~H~M~2~1fixatives_in_an_attempt_to_find_one_that_would_give_us_the~Kbr_~H~M~2~1cellular_morphology_that_we_were_used_to_and_also_be_optimal~Kbr_~H~M~2~1for_IHC~HICC._We_tested_out_the_following_fixatives~I~Kbr_~H~M~2~1~Kbr_~H~M~2~110~7_NBF~Kbr_~H~M~2~170~7_EtOH~Kbr_~H~M~2~170~7_MeOH~Kbr_~H~M~2~1Carnoy~9s~Kbr_~H~M~2~1methacarn_~ACarnoy~9s_with_methanol_instead_of_ethanol~B~Kbr_~H~M~2~1zinc_formalin~E_unbuffered~Kbr_~H~M~2~1buffered_zinc_formalin~Kbr_~H~M~2~1~Kbr_~H~M~2~1The_10~7_NBF_of_course_gave_us_the_morphology_we_were_used_to~E~Kbr_~H~M~2~1and_if_fixation_times_were_kept_to_24~F48_hours~E_epitope~Kbr_~H~M~2~1retrieval_was_not_required_for_most_antibodies._If_tissues~Kbr_~H~M~2~1needed_to_be_stored_longer_than_48_hours~E_they_were_stored_in~Kbr_~H~M~2~170~7_EtOH_until_ready_to_be_processed._Of_all_the_fixatives_we~Kbr_~H~M~2~1tested~E_the_worst_for_IHC_was_70~7_EtOH~E_then_Carnoy~9s._The~Kbr_~H~M~2~1best_for_IHC_was_70~7_MeOH._Cellular_morphology_for_both_of~Kbr_~H~M~2~1these_was_not_all_that_great._Methacarn_gave_us_both_good~Kbr_~H~M~2~1morphology_and_good_IHC._The_zinc_formalins_gave_excellent~Kbr_~H~M~2~1morphology_in_many_organs~E_and_good_IHC_staining._It_should_be~Kbr_~H~M~2~1noted_that_the_zinc_formalins_have_difficulty_penetrating_the~Kbr_~H~M~2~1hematopoietic_organs_as_they_react_more_with_the_RBCs_and~Kbr_~H~M~2~1therefore_penetration_is_much_slower._As_those_are_the_organs~Kbr_~H~M~2~1of_interest_in_our_laboratory~E_we_use_standard_NBF.~Kbr_~H~M~2~1~Kbr_~H~M~2~1We_have_found_that_if_the_tissues_are_trimmed_to_a_thickness_of~Kbr_~H~M~2~1no_more_than_3_mm_prior_to_immersion_in_NBF~E_fixative_solutions~Kbr_~H~M~2~1are_changed_at_1_and_12_hours~E_and_after_24_hours_in_fixative~Kbr_~H~M~2~1are_transferred_to_70~7_EtOH~E_both_cellular_morphology_and_IHC~Kbr_~H~M~2~1staining_are_excellent.~Kbr_~H~M~2~1~Kbr_~H~M~2~1One_of_these_days_when_I_have_some_time_I~9d_like_to_try~Kbr_~H~M~2~1some_of_the_other_fixatives~E_as_well_as_some_of_the_commercial~Kbr_~H~M~2~1ones_that_are_out_there~E_just_to_see_what_the_total_comparisons~Kbr_~H~M~2~1are_going_to_be_like._I_would_also_like_to_note_that_Bouin~9s~Kbr_~H~M~2~1has_seemed_to_work_pretty_much_all_right_as_I_have_been_doing~Kbr_~H~M~2~1IHC_on_some_Bouin~9s_fixed_testes_lately_without_problems.~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Perfusion_fixative_for_electron_microscopy~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1What_is_a_suitable_fixative_for_vascular_perfusion_of_rats~E~Kbr_~H~M~2~1and_subsequent_electron_microscopy_of_tissues~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1A_neutral~E_buffered~E_isotonic_formaldehyde~Fglutaraldehyde~Kbr_~H~M~2~1mixture_should_be_fine_for_any_kind_of_electron_microscopy.~Kbr_~H~M~2~1Many_workers_like_to_use_paraformaldehyde_as_the_source~Kbr_~H~M~2~1of_formaldehyde.~Kbr_~H~M~2~1~Kbr_~H~M~2~1A_classical_mixture_is_M._J._Karnovsky~9s_~AJ._Cell_Biol.~Kbr_~H~M~2~127~I_137A~F138A~E_1965~B._This_is_probably_the_most~Fcited~Kbr_~H~M~2~1unrefereed_abstract~3_It_contains_approximately_4~7_formaldehyde~Kbr_~H~M~2~1and_5~7_glutaraldehyde_in_approximately_0.1_M_phosphate_or~Kbr_~H~M~2~1cacodylate_buffer._Final_pH_~L_7.2._If_cacodylate_~Atoxic~3~B~Kbr_~H~M~2~1is_used~E_add_calcium_chloride_~A0.5_mg~Hml~B_to_improve~Kbr_~H~M~2~1preservation_of_membrane_phospholipids.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Probably_this_fixative_is_frequently_misquoted~E_and_the~Kbr_~H~M~2~1literature_is_full_of_references_to_~4half~Fstrength~Kbr_~H~M~2~1Karnovsky~E~4_which_probably_means_half_the_glutaraldehyde~Kbr_~H~M~2~1concentration._A_glutaraldehyde_concentration_of_1_to_2~7~Kbr_~H~M~2~1is_commonly_considered_adequate_in_mixtures_of_this~Kbr_~H~M~2~1kind.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Fixation_of_frozen_sections.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1What_is_the_best_fixative_for_frozen_sections~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Unfixed_tissue~E_cut_with_a_cryostat_~Athin_sections~B_or_a~Kbr_~H~M~2~1vibrating_microtome_~Athick_sections~B_should_be_fixed_if_this~Kbr_~H~M~2~1is_compatible_with_the_staining_technique_to_be_used.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Many_enzyme_histochemical_methods_demand_unfixed_sections~E~Kbr_~H~M~2~1and_so_do_immunohistochemical_methods_with_some_~Afortunately~Kbr_~H~M~2~1not_most~B_primary_antibodies._Enzyme_incubations_are_often~Kbr_~H~M~2~1terminated_by_moving_the_slide_or_coverslip_bearing_the~Kbr_~H~M~2~1cryostat_section_from_the_incubation_medium_into_a_neutral~E~Kbr_~H~M~2~1buffered_formaldehyde_fixative.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Even_~4minimal~4_~A~L_inadequate~B_fixation_before_staining_will~Kbr_~H~M~2~1greatly_improve_the_structural_preservation_of_tissue._Many~Kbr_~H~M~2~1enzymes_will_survive_either_a_minute_or_two_in_neutral~E~Kbr_~H~M~2~1buffered_formaldehyde~E_followed_by_a_wash_in_buffered_saline.~Kbr_~H~M~2~1Some_enzymes_and_most_antigens_will_survive_immersion_of_the~Kbr_~H~M~2~1slide_or_coverslip_in_cold_~Aabout_0_C~B_acetone_for_half_a~Kbr_~H~M~2~1minute._The_acetone_is_allowed_to_evaporate_before_immersing~Kbr_~H~M~2~1the_section_in_incubation_medium.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Cryostat_sections_may_also_be_fixed_by_heating~E_but_this~Kbr_~H~M~2~1inactivates_most_enzymes._A_drop_of_an_ethanol~Fpoly~Aethylene~Kbr_~H~M~2~1glycol~B_mixture_is_placed_on_the_section_and_the_temperature~Kbr_~H~M~2~1brought_up_to_55_C_in_a_microwave_oven._~AA_special_laboratory~Kbr_~H~M~2~1oven_is_needed_to_get_this_amount_of_control.~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1References.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Kiernan_JA_1999._Histological_and_histochemical_Methods~E_3rd~Kbr_~H~M~2~1ed._Oxford~I_Butterworth~FHeinemann.~Kbr_~H~M~2~1Kok_LP_~8amp~J_Boon_ME_1992._Microwave_Cookbook_for_Microscopists.~Kbr_~H~M~2~1Leiden~I_Coulomb_Press.~Kbr_~H~M~2~1Pearse_AGE_1980._Histochemistry~E_4th_ed._Vol_1.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Non~Fformaldehyde_commercial_fixatives~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Commercially_available_fixatives_are_touted_variously_as~Kbr_~H~M~2~1~4non~Fcrosslinking~E~4_~4less~Fcrosslinking~E~4_~4formaldehyde~Ffree~E~4~Kbr_~H~M~2~1~4better_for_immunohistochemistry~E~4_~4less_toxic~E~4_~Eetc.~E_etc.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Is_there_a_recent_review~E_or_can_someone_share_a_list_of~Kbr_~H~M~2~1names_of_commercially_available_fixatives_~Asupposedly_better~Kbr_~H~M~2~1for_immunohistochemistry~B_and_their_vendors~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Here_are_all_of_the_ones_that_I_know_about~J_some_of_them_may_be~Kbr_~H~M~2~1sold_under_different_names_by_other_vendors~I~Kbr_~H~M~2~1~Kbr_~H~M~2~1GlyoFix_from_Shandon_Lipshaw_uses_glyoxal_as_the_active~Kbr_~H~M~2~1ingredient~J_produces_aldehyde~Ftype_fixation_patterns.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Histochoice_from_Amresco~J_active_ingredients_essentially~Kbr_~H~M~2~1undisclosed_~Aaldehydic_addition_compounds~B~J_mode_of_action~Kbr_~H~M~2~1unknown.~Kbr_~H~M~2~1~Kbr_~H~M~2~1HistoFix~E_formerly_from_Trend_Scientific~E_perhaps_still~Kbr_~H~M~2~1available_from_Baxter~E_contains_pyrrolid~F2~Fone~E_a_polyol~E~Kbr_~H~M~2~1a_urea_and_a_zinc_salt~J_mode_of_action_unknown.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Mirsky~9s_Fixative_from_National_Diagnostics_is_an_aqueous~Kbr_~H~M~2~1solution_of_a_complex_di~Faldehyde_~Apossible_di~Faldehyde~Kbr_~H~M~2~1starch~B~J_mode_of_action_may_be_aldehyde~Flike~E_but_very_slow.~Kbr_~H~M~2~1~Kbr_~H~M~2~1NoToX_from_EarthSafe_Industries~E_uses_a_complex_aldehyde~Kbr_~H~M~2~1~Apossibly_di~Faldehyde_glucose~B_in_about_70~7_alcohol_with~Kbr_~H~M~2~1antiseptic_and_antifungal_agents~J_produces_a_combination_of~Kbr_~H~M~2~1aldehyde~F_and_alcohol~Ftype_fixation_patterns.~Kbr_~H~M~2~1~Kbr_~H~M~2~1OmniFix_II_and_OmniFix_2000_from_AnCon_Genetics_is_an~Kbr_~H~M~2~1alcohol~Fbased_solution_containing_glycol_and_salts~J~Kbr_~H~M~2~1produces_alcohol~Ftype_fixation_patterns.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Prefer_from_Anatech_Ltd.~E_uses_glyoxal_as_the_active~Kbr_~H~M~2~1ingredient~J_produces_aldehyde~Ftype_fixation_patterns.~Kbr_~H~M~2~1~Kbr_~H~M~2~1SafeFix_II_from_CMS_uses_glyoxal_as_the_active_ingredient~J~Kbr_~H~M~2~1produces_aldehyde~Ftype_fixation_patterns.~Kbr_~H~M~2~1~Kbr_~H~M~2~1STF_~AStreck_Tissue_Fixative~B_from_Streck_Laboratories_contains~Kbr_~H~M~2~1diazolidinyl_urea~E_2~Fbromo~F2~Fnitropropane~F1~E3_diol~E_zinc~Kbr_~H~M~2~1sulfate_and_a_small_amount_of_formaldehyde_as_active~Kbr_~H~M~2~1ingredients~J_mode_of_action_unknown.~Kbr_~H~M~2~1~Kbr_~H~M~2~1There_are_two_fixatives_intended_for_microwave_use~I~Kbr_~H~M~2~1~Kbr_~H~M~2~1Preserve_from_Energy_Beam_Sciences_uses_glyoxal_as_the_active~Kbr_~H~M~2~1ingredient~J_produces_aldehyde~Ftype_fixation_patterns.~Kbr_~H~M~2~1~Kbr_~H~M~2~1MicroFix_from_Energy_Beam_Sciences_is_an_alcohol~Hpolyethylene~Kbr_~H~M~2~1glycol_solution._It_replaces_Merck~9s_KryoFix~E_which_is_no~Kbr_~H~M~2~1longer_available~J_produces_alcohol~Ftype_fixation_patterns.~Kbr_~H~M~2~1~Kbr_~H~M~2~1A_rather_uncomplimentary_comparison_of_some_of_these_products~Kbr_~H~M~2~1~AHistochoice~E_KryoFix~E_Mirsky~E_NoToX~E_Omnifix_II_and_STF~B~Kbr_~H~M~2~1has_been_published_~APrento_~8amp~J_Lyon~E_1997._Commercial_formalin~Kbr_~H~M~2~1substitutes_for_histopathology._Biotechnic_~8amp~J_Histochemistry~E~Kbr_~H~M~2~172~I273~F282~B._Readers_should_note_that_none_of_them_were_used~Kbr_~H~M~2~1as_directed_or_intended_by_the_manufacturers_~Afixation_at_4~Kbr_~H~M~2~1degrees_C~B~E_so_the_results_are_questionable._Also~E_none_of_the~Kbr_~H~M~2~1glyoxal~Fbased_fixatives_~AGlyoFix~E_Prefer~E_SafeFix_II~E_Preserve~B~Kbr_~H~M~2~1were_tested~J_these_seem_to_be_the_most_favored_substitutes_in~Kbr_~H~M~2~1the_USA_at_least~E_because_they_most_nearly_mimic_the~Kbr_~H~M~2~1morphological_patterns_obtained_with_formalin_without~Kbr_~H~M~2~1formaldehyde~9s_unfavorable_effects_on_immunoreactivity.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Glutaraldehyde_and_immunohistochemistry~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Does_glutaraldehyde_fixative_~A4~7_paraformaldehyde~E~Kbr_~H~M~2~10.5~7_glutaraldehyde~B_interfere_with_fluorescent~Kbr_~H~M~2~1immunohistochemistry~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_1.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Glutaraldehyde~E_because_of_its_reactivity_and_speed~E_can~Kbr_~H~M~2~1seriously_interfere_with_antibody_binding_and_lectin_binding~Kbr_~H~M~2~1causing_considerable_non~Fspecific_binding._It_is_also_difficult~Kbr_~H~M~2~1to_remove_excess_glutaraldehyde_from_tissue_components._I_would~Kbr_~H~M~2~1not_recommend_it~9s_use_for_such_studies~E_as_in_my_hands_the~Kbr_~H~M~2~1results_have_been_inconsistent.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Barry_Rittman~Kbr_~H~M~2~1~Abrittman~PAT~Qmail.db.uth.tmc.edu~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_2.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Tissues_fixed_in_glutaraldehyde_exhibit_increased_autofluorescence~E~Kbr_~H~M~2~1which_is_probably_due_to_glutaraldehyde~Famino_acid_compounds_that~Kbr_~H~M~2~1are_formed_as_part_of_the_fixative_action._Glutaraldehyde_also~Kbr_~H~M~2~1introduces_free_aldehyde_groups_into_the_tissue~E_and_these_will~Kbr_~H~M~2~1bind_any_protein_reagents_that_are_applied._The_nonspecific~Kbr_~H~M~2~1binding_of_antibodies_can_be_reduced_by_pretreatment_with_a~Kbr_~H~M~2~1blocking_protein_~Asuch_as_bovine_albumin~E_or_serum_from_the~Kbr_~H~M~2~1species_in_which_a_secondary_antibody_was_raised~B._Before_the~Kbr_~H~M~2~1blocking_treatment_it_is_advisable_to_do_a_chemical_aldehyde~Kbr_~H~M~2~1blockade_~AHistochemistry_textbooks_contain_several_methods~B.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Isopentane~I_alternative_names~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Is_isopentane_the_same_as_2~Fmethyl_butane~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Yes._It_is_also_known_as_ethyldimethylmethane~Kbr_~H~M~2~1All_are_~ACH3~B2CHCH2CH3.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Anita_Jennings~Kbr_~H~M~2~1~Ajennings~PAT~Qmayo.edu~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1Back_to_Table_of_Contents~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Lidocaine_in_perfusion_fixation~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Lidocaine_can_be_added_to_the_fixative_during_perfusion_I_would~Kbr_~H~M~2~1appreciate_hearing_the_Lidocaine_concentration_again.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1This_is_the_recipe_for_lidocaine_I_used_for_perfusion~Ffixing~Kbr_~H~M~2~1mormyrids_~Aan_electric_fish~B~I~Kbr_~H~M~2~1~Kbr_~H~M~2~1Lidocaine_~A~L_lignocaine_~L_xylocaine~B_for_use_in_perfusion~Kbr_~H~M~2~1fixation_~AUsed_to_relax_blood_vessels_to_permit_more_complete~Kbr_~H~M~2~1exchange_~8amp~J_infiltration_of_fixative.~B~I~Kbr_~H~M~2~1~Kbr_~H~M~2~1Lidocaine_50_mg_per_mL~E_dissolved_95~7_EtOH_~Awarm_to_dissolve~B.~Kbr_~H~M~2~1Add_slowly_to_perfusion_solution_with_stirring_to_make_final~Kbr_~H~M~2~1lidocaine_solution_concentration_wanted_~Ae.g._1mg~HmL_~L_0.1~7~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1Note~I_Do_not_add_the_lidocaine_directly_to_the_perfusion~Kbr_~H~M~2~1solution~E_especially_if_the_solution_contains_salts~3_The~Kbr_~H~M~2~1lidocaine_will_not_go_into_solution.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Philip_Oshel~Kbr_~H~M~2~1~Aoshel~PAT~Qshout.net~Kbr_~H~M~2~1or_poshel~PAT~Qhotmail.com~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1Back_to_Table_of_Contents~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Michel~9s_fluid_for_transporting_cells_or_specimens~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Does_anyone_have_any_references_for_Michel~9s_Fixative_or_Fluid~N~Kbr_~H~M~2~1We_use_it_for_an_immunofluorescence_holding_medium~E_but_I_don~9t~Kbr_~H~M~2~1have_a_reference_on_it_for_the_manual_..._~Aand_I_would_like_to~Kbr_~H~M~2~1read_about_it_just_for_my_own_knowledge~B.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Here~9s_my_procedure_sheet_for_Michel~9s_transport_medium.~Kbr_~H~M~2~1~Kbr_~H~M~2~1MICHEL~9S_TRANSPORT_MEDIUM~Kbr_~H~M~2~1~Kbr_~H~M~2~1Michel~9s_transport_medium_~Apronounced_mee~FSHELL~B_is_used_to~Kbr_~H~M~2~1transport_specimens_~Asuch_as_renal_biopsies_and_lymph_nodes~B_for~Kbr_~H~M~2~1immunofluorescence_studies._It_is_not_a_fixative~E_and_is_not~Kbr_~H~M~2~1suitable_for_any_other_use_~Aparticularly~E_it_is_not_suitable_for~Kbr_~H~M~2~1transporting_living_cells_for_flow_cytometry~B._It_should_be~Kbr_~H~M~2~1stored_refrigerated_~Anot_frozen~B_until_use._Specimens_may_be~Kbr_~H~M~2~1kept_in_it_at_room_temperature_until_they_can_be_delivered_to~Kbr_~H~M~2~1the_reference_laboratory._Zeus_Medium~E_a_commercial_product~E_is~Kbr_~H~M~2~1probably_similar.~Kbr_~H~M~2~1~Kbr_~H~M~2~11.0_M_potassium_citrate_buffer_pH_7.0~I~Kbr_~H~M~2~1dissolve_21.0_g_citric_acid_monohydrate~Kbr_~H~M~2~1~Aor_19.2_g_citric_acid_anhydrous~B~Kbr_~H~M~2~1in_30_mL_of_hot_deionized_or_distilled_water._Cool.~Kbr_~H~M~2~1Adjust_pH_to_7.0_with_1_M_potassium_hydroxide_~Aabout_35_mL~B~Kbr_~H~M~2~1Dilute_to_100_mL_with_more_water.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Washing_solution~I~Kbr_~H~M~2~1~Kbr_~H~M~2~125_mL_1.0_M_potassium_citrate_buffer~Kbr_~H~M~2~150_mL_0.1_M_magnesium_sulfate_heptahydrate_~AF.W._246.5~B~Kbr_~H~M~2~150_mL_0.1_M_N~Fethyl_maleimide_~A~L_12.5_g_in_1_L_of_water~B~Kbr_~H~M~2~1~ASigma_E3876.~B~Kbr_~H~M~2~1Water_to_make_1_L~Kbr_~H~M~2~1Adjust_to_pH_7.0_with_1_M_potassium_hydroxide~Kbr_~H~M~2~1Store_in_refrigerator._~ACost_about_$50~H25_g_in_1994.~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1Transport_medium~I~Kbr_~H~M~2~1~Kbr_~H~M~2~1Dissolve_55_grams_of_ammonium_sulfate_in_100_mL_washing~Kbr_~H~M~2~1solution._~AAdd_slowly~E_with_mechanical_stirring.~B~Kbr_~H~M~2~1Adjust_pH_to_about_6.9_with_1_M_potassium_hydroxide~Kbr_~H~M~2~1~A~8lt~J_2_mL_needed~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1Specimens_can_be_held_at_room_temperature_for_five_days_in~Kbr_~H~M~2~1transport_medium_before_processing._Specimens_received_in~Kbr_~H~M~2~1transport_medium_should_be_washed_in_three_changes_of_washing~Kbr_~H~M~2~1solution~E_10_minutes_each_wash.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Reference~I_Michel_B._Milner_Y._David_K._Preservation_of~Kbr_~H~M~2~1tissue~Ffixed_immunoglobulins_in_skin_biopsies_of_patients_with~Kbr_~H~M~2~1lupus_erythematosus_and_bullous_diseases._A_preliminary_report.~Kbr_~H~M~2~1J._Invest._Dermatol._59~I_49~F452_~A1973~B.~Kbr_~H~M~2~1~Kbr_~H~M~2~1This_procedure_received_from_J._Charles_Jennette_MD~E_Immunopathology~Kbr_~H~M~2~1Laboratory~E_North_Carolina_Memorial_Hospital~E_Chapel_Hill_NC_27514~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Can_someone_experienced_with_a_microwave_processor_give_advice~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_1.~Kbr_~H~M~2~1~Kbr_~H~M~2~1In_making_your_final_decision_about_the_purchase_of_a_laboratory~Kbr_~H~M~2~1microwave_oven~E_you_may_also_find_it_helpful_to_use_some_simple~Kbr_~H~M~2~1microwave_calibration_tools_to_determine_objectively_if_a~Kbr_~H~M~2~1particular_microwave_oven_will_suit_your_specific_needs.~Kbr_~H~M~2~1~Kbr_~H~M~2~1These_tools_are_quick_and_simple_assessments_that_show_you_just~Kbr_~H~M~2~1how_evenly_your_clinical_specimens_will_be_heated_in_a_microwave~Kbr_~H~M~2~1oven.~Kbr_~H~M~2~1~Kbr_~H~M~2~11._Neon_Bulb_Array.~Kbr_~H~M~2~1Because_our_eyes_can_not_sense_microwaves~E_they_appear~Kbr_~H~M~2~1invisible_to_us._A_Neon_Bulb_Array_is_a_tool_that_indirectly~Kbr_~H~M~2~1shows_the_nonuniformity_of_microwave_power_in_a_microwave~Kbr_~H~M~2~1oven._In_principle~E_microwave_irradiation_increases_the~Kbr_~H~M~2~1kinetic_energy_of_the_neon_gas_molecules._The_neon_bulbs~Kbr_~H~M~2~1glow_orange_where_the_microwave_power_is_high_enough_to~Kbr_~H~M~2~1ionize_the_gas_molecules_~A~X5_mw~Hcm2~B._The_neon_bulb_array_is~Kbr_~H~M~2~1useful_for_determining_the_areas_of_uniform_power~E_cycle~Kbr_~H~M~2~1time~E_and_magnetron_warm~Fup_time_in_a_microwave_oven~Kbr_~H~M~2~1~Kbr_~H~M~2~12._The_Agar~FSaline~FGiemsa_tissue_phantom.~Kbr_~H~M~2~1Agar~FSaline~FGiemsa_tissue_phantoms_are_used_to_simulate_the~Kbr_~H~M~2~1size~E_shape~E_and_absorbance_characteristics_of_biological~Kbr_~H~M~2~1specimens_to_verify_that_the_microwave_oven_will_uniformly~Kbr_~H~M~2~1heat_the_specimens.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Small_agar_phantoms_~A1_cm_x_0.5_cm2_blocks_or_2_cm_diameter~Kbr_~H~M~2~1by_0.3_cm_thickness_discs~B_that_contain_0.002~7_commercial~Kbr_~H~M~2~1Giemsa_stain_are_added_to_molten_2~7_agar_in_0.9~7_sodium~Kbr_~H~M~2~1chloride._The_Giemsa_dyes_respond_to_microwave_heating_by~Kbr_~H~M~2~1showing_different_colors_at_different_temperatures._When_ASG~Kbr_~H~M~2~1tissue_phantoms_are_irradiated_in_an_optimized_microwave~Kbr_~H~M~2~1cavity~E_they_show_a_uniform_color_change.~Kbr_~H~M~2~1~Kbr_~H~M~2~1These_tools_have_been_described_and_published_in_peer~Freviewed~Kbr_~H~M~2~1journals_since_1990_and_have_been_independently_verified_by~Kbr_~H~M~2~1other_laboratories._They_are_commercially_available_or_you_can~Kbr_~H~M~2~1prepare_them_yourself.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Brief_list_of_references~Kbr_~H~M~2~1~Kbr_~H~M~2~11._Login~E_G._R.~E_N._Tanda~E_and_A._M._Dvorak._Calibrating_and~Kbr_~H~M~2~1standardizing_microwave_ovens_for_microwave~Faccelerated_specimen~Kbr_~H~M~2~1preparation._A_review._Cell_Vision_3~I_172~F179~E_1996.~Kbr_~H~M~2~12._Login~E_G._R.~E_and_A._M._Dvorak._The_Microwave_Toolbook._A_Practical~Kbr_~H~M~2~1Guide_for_Microscopists._Boston~I_Beth_Israel_Hospital~E_1994.~Kbr_~H~M~2~13._Login~E_G._R.~E_J._B._Leonard~E_and_A._M._Dvorak._Calibration_and~Kbr_~H~M~2~1standardization_of_microwave_ovens_for_fixation_of_brain_and~Kbr_~H~M~2~1peripheral_nerve_tissue._Companion_to_Methods_Enzymol_15~I_~Ain~Kbr_~H~M~2~1press~B~E_1998.~Kbr_~H~M~2~14._Login~E_G._R._The_need_for_clinical_laboratory_standards_for~Kbr_~H~M~2~1microwave~Faccelerated_procedures._J_Histotechnol_21~I_1~F3~E_1998~Kbr_~H~M~2~1~AEditorial~B.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_2.~Kbr_~H~M~2~1~Kbr_~H~M~2~1My_experience_thus_far_is_purely_from_a_vendors_view.~Kbr_~H~M~2~1The_benefits_so_far~I~Kbr_~H~M~2~1~Kbr_~H~M~2~11._You_can_process_without_xylene~Kbr_~H~M~2~12._Turnaround_can_be_minutes_as_opposed_to_hours.~Kbr_~H~M~2~13._cost_savings_about_1~H5_of_a_traditional_processor.~Kbr_~H~M~2~1~ANot_counting_the_reagent_savings~B~Kbr_~H~M~2~14._Loads_of_up_to_90_cassettes_can_be_processed_in_one_run.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Paraformaldehyde~I_why_won~9t_it_dissolve~N~Kbr_~H~M~2~1~AAnswer_includes_other_information_about_formaldehyde_and_fixation~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Why_will_paraformaldehyde_not_dissolve_in_unaltered_seawater~Kbr_~H~M~2~1without_added_sodium_hydroxide~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Paraformaldehyde_is_a_white_solid_formed_by_combination_of_large~Kbr_~H~M~2~1numbers_of_formaldehyde_molecules_in_an_aqueous_solution~I_a~Kbr_~H~M~2~1polymer._Formaldehyde~E_HCHO~E_is_a_gas_and_strictly_speaking_it~Kbr_~H~M~2~1doesn~9t_exist_in_aqueous_solution_because_it_tacks_on_a_water~Kbr_~H~M~2~1molecule_to_form_methylene_hydrate~E_which_is_HO~FCH2~FOH._This_is~Kbr_~H~M~2~1the_active_ingredient_of_fixatives._Methylene_hydrate_molecules~Kbr_~H~M~2~1just_love_one_another~E_and_join_together_~Aeliminating_H2O~E_so_I~Kbr_~H~M~2~1suppose_it~9s_really_the_original_formaldehyde_carbon_atoms_that~Kbr_~H~M~2~1are_so_affectionate~B_to_make_polymers_of_all_sizes._In_commercial~Kbr_~H~M~2~1formalin_~A37~F40~7_HCHO_by_weight~B_the_polymer_molecules_are_small~Kbr_~H~M~2~1enough_to_stay_in_solution._In_paraformaldehyde_they_are_big~Kbr_~H~M~2~1enough_to_be_insoluble.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Manufacturers_add_some_methanol_to_formalin._This_retards_the~Kbr_~H~M~2~1formation_of_large_polymer_molecules_~Asee_Recommended_Reading_if~Kbr_~H~M~2~1you_want_to_know_why~B._Probably_the_methanol_doesn~9t_affect~Kbr_~H~M~2~1fixative_properties_when_diluted~E_though_some_people_in_the_late~Kbr_~H~M~2~11950s_claimed_that_it_did._If_you_buy_paraformaldehyde~E_you_can~Kbr_~H~M~2~1depolymerize_it_yourself_and_get_a_solution_of_~4formaldehyde~4~Kbr_~H~M~2~1~Aactually_methylene_hydrate~B_that_doesn~9t_contain_any_methanol.~Kbr_~H~M~2~1~Kbr_~H~M~2~1From_what_I~9ve_said_so_far~E_~SPlease_Take_Note~3~S_it_follows_that~Kbr_~H~M~2~1there_is_no_such_thing_as_a_~42~7_~Aor_any_other_~7~B_paraformaldehyde~4~Kbr_~H~M~2~1solution._Paraformaldehyde_is_a_high_polymer~E_and_its_molecules~Kbr_~H~M~2~1are_too_big_to_dissolve_in_water~E_alcohol_or_anything_else.~Kbr_~H~M~2~1~Kbr_~H~M~2~1You_have_to_depolymerize_paraformaldehyde_to_get_it_to_~4dissolve~4~Kbr_~H~M~2~1and_form_a_formaldehyde_~Areally_methylene_hydrate~B_solution._The~Kbr_~H~M~2~1depolymerization_is_a_reaction_of_the_polymer_with_water~I_a~Kbr_~H~M~2~1hydrolysis._It_needs_hydroxide_ions_~AOH~F~B_as_a_catalyst~E_and_also~Kbr_~H~M~2~1some_heat_to_get_the_job_done_in_reasonable_time._In_the_making_of~Kbr_~H~M~2~1ordinary_phosphate~Fbuffered_formaldehyde_from_paraformaldehyde~E~Kbr_~H~M~2~1the_usual_procedure_is_to_heat_the_PF_with_the_dibasic_sodium~Kbr_~H~M~2~1phosphate_component_of_the_buffer._This_contains_enough_OH~F_ions~Kbr_~H~M~2~1to_catalyse_the_hydrolysis_and_depolymerization._You_add_the~Kbr_~H~M~2~1acidic_part_of_the_buffer_~Asodium_or_potassium_dihydrogen~Kbr_~H~M~2~1phosphate~B_when_the_solution_has_become_transparent._This_occurs~Kbr_~H~M~2~1when_the_temperature_reaches_about_60_C._It_should_not_be~Kbr_~H~M~2~1necessary_to_go_any_hotter_than_that.~Kbr_~H~M~2~1~Kbr_~H~M~2~1In_the_earliest_recommended_fixatives_that_started_with~Kbr_~H~M~2~1paraformaldehyde~E_a_few_drops_of_sodium_hydroxide_were_added_to_a~Kbr_~H~M~2~1heated_suspension_of_paraformaldehyde_in_water_or_saline._This~Kbr_~H~M~2~1hardly_affected_the_pH_of_the_final_solution.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Additional_question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1My_supervisor_~Awho_has_been_trained_in_histology~E_unlike_myself~3~3~B_said~Kbr_~H~M~2~1that_in_most_of_my_staining_and_fixative_methods_can_have_the_phosphate~Kbr_~H~M~2~1buffer_component_replaced_by_seawater_with_no_problems_as_seawater_is_a~Kbr_~H~M~2~1buffer~E_at_the_right_osmolarity_for_fish_tissue._Is_this_the_case~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer~E_continued.~Kbr_~H~M~2~1~Kbr_~H~M~2~1I_don~9t_know_how_good_a_buffer_sea_water_is~E_but_it~9s_unlikely~Kbr_~H~M~2~1to_be_as_robust_as_0.1M_phosphate._In_a_fixative_the_osmolarity~Kbr_~H~M~2~1is_more_important_than_the_pH~E_but_for_a_slowly_acting_agent~Kbr_~H~M~2~1like_formaldehyde_or_a_slowly_penetrating_one_like_osmium~Kbr_~H~M~2~1tetroxide~E_the_solvent_should_be_as_similar_as_possible_to_the~Kbr_~H~M~2~1extracellular_fluids_of_whatever_you~9re_fixing._If_the~Kbr_~H~M~2~1formaldehyde_~Atakes_hours_to_do_its_stuff~B_is_mixed_with_more~Kbr_~H~M~2~1rapidly_acting_fixative_agents_~Aalcohol~E_mercuric_chloride~E~Kbr_~H~M~2~1picric_acid_etc.~E_which_act_as_soon_as_they_reach_the_cells~B~E~Kbr_~H~M~2~1the_osmolarity_is_less_important~E_and_most_such_mixtures_are~Kbr_~H~M~2~1acidic_too._The_formaldehyde_does_its_cross~Flinking_after_the~Kbr_~H~M~2~1proteins_have_been_insolubilized_by_the_coagulant_components.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Readings~I~Kbr_~H~M~2~1For_formaldehyde_chemistry~I_Walker~E_JF_1964.~Kbr_~H~M~2~1Formaldehyde._2nd_ed._New_York~I_Reinhold~J_London~I_Chapman~Kbr_~H~M~2~1~8amp~J_Hall.~Kbr_~H~M~2~1For_how_formaldehyde_works~I_Pearse~E_AGE~I~Kbr_~H~M~2~1Histochemistry~E_Theoretical_and_Applied._Any_edition_of_this~Kbr_~H~M~2~1book_should_be_OK._There~9s_also_lots_of_erudite_discussion~Kbr_~H~M~2~1in_Baker~E_JR_~A1958~B_Principles_of_Biological_Microtechnique.~Kbr_~H~M~2~1London~I_Methuen~E_which_is_a_great_classic_in_the_field.~Kbr_~H~M~2~1For_some_stuff_on_the_slowness_of_formaldehyde~Kbr_~H~M~2~1fixation_and_importance_of_an_isotonic_buffer~I_Paljarvi~EL~E~Kbr_~H~M~2~1Garcia~EJH_~8amp~J_Kalimo~EH_1979._Histochem._J._11~E_267~F269~J~Kbr_~H~M~2~1Schook~E_P_1980._Acta_morph._Neerl.~FScand._18~I_31~F45._See~Kbr_~H~M~2~1also_some_of_MA_Hayat~9s_books_on_techniques_for_electron~Kbr_~H~M~2~1microscopy~E_which_discuss_the_subject_thoroughly.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Saccomano~9s_fixative~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Does_anyone_have_a_recipe_for_Saccomanno_fixative_~Aa_cytology~Kbr_~H~M~2~1fixative~B_which_gives_the_molecular_weight_of_the_Carbowax~Kbr_~H~M~2~1~Apolyethylene_glycol~B_in_the_solution~N_Thanks_in_advance~3~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1This_formula_is_from_Koss._Roughly_equal_volumes_of~Kbr_~H~M~2~1Saccomanno~9s_fixative_can_be_added_to_liquid_cytologic~Kbr_~H~M~2~1specimens_such_as_sputum~E_urine~E_bronchial_washings~E_and~Kbr_~H~M~2~1pleural_and_peritoneal_fluids_to_stabilize_them_at_room~Kbr_~H~M~2~1temperature_until_they_can_be_prepared_as_filter_or~Kbr_~H~M~2~1cytocentrifuge_preparations_or_cell_blocks~E_and_it_also_works~Kbr_~H~M~2~1fairly_well_for_small_biopsy_specimens._It_is_not_suitable_for~Kbr_~H~M~2~1ThinPrep_preparations~E_for_which_a_special_fixative_is~Kbr_~H~M~2~1required.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Saccomanno~9s_fixative_is_50~7_alcohol_which_contains~Kbr_~H~M~2~1approximately_2~7_of_Carbowax_1540_~AUnion_Carbide_Corporation~E~Kbr_~H~M~2~1UCAR~B._Carbowax_1540_is_solid_at_room_temperature~E_with_a~Kbr_~H~M~2~1melting_point_of_43_to_46_C._To_avoid_having_to_melt_it~Kbr_~H~M~2~1whenever_the_fixative_is_prepared~E_a_stock_solution_can_be~Kbr_~H~M~2~1propared_by_melting_of_Carbowax_~Amelted_in_an_incubator_or_hot~Kbr_~H~M~2~1air_oven_at_50_to_100_C~B_and_adding_it_to_an_equal_volume_of~Kbr_~H~M~2~1water_or_50~7_alcohol._The_mixture_will_not_solidify.~Kbr_~H~M~2~1Saccomanno~9s_fixative_can_then_be_prepared_with_430_mL_of~Kbr_~H~M~2~1water~E_530_mL_of_95~7_ethanol~E_and_40_mL_of_the_stock_Carbowax~Kbr_~H~M~2~1solution._Some_light_green_SF_or_fast_green_FCF_can_be_added~Kbr_~H~M~2~1to_color_the_fixative._Koss_warns_that_the_denaturants_in~Kbr_~H~M~2~1reagent_alcohol_may_cause_excessive_hardening_of_mucus.~Kbr_~H~M~2~1~Kbr_~H~M~2~1I_suppose_that_the_1540_is_the_molecular_weight~E_but_basically~Kbr_~H~M~2~1it~9s_a_catalog_number_for_a_long_series_of_these_UCAR_products~Kbr_~H~M~2~1that_range_from_thin_liquids_to_dense_paraffin~Flike_waxes.~Kbr_~H~M~2~1~Kbr_~H~M~2~1From_Leopold_G._Koss~E_Diagnostic_Cytology_and_Its_Histologic~Kbr_~H~M~2~1Bases~E_3rd_ed.~E_Lippincott_1979~E_page_1192_I_don~9t_have_the~Kbr_~H~M~2~1current_edition_of_this_venerable_tome._I_have_never_tried_to~Kbr_~H~M~2~1make_Saccomanno~9s_fixative~E_but_those_who_have_rank_it_right~Kbr_~H~M~2~1up_there_with_hanging_wallpaper_as_a_good_way_to_wind_up~Kbr_~H~M~2~1screaming.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Zinc~Fcontaining_fixatives~I_What_has_been_published~N~Kbr_~H~M~2~1~AAnswers_include_references~E_opinions_and_discussion.~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1Questions.~Kbr_~H~M~2~1~Kbr_~H~M~2~1What_published_work_is_available_with_evaluations_of~Kbr_~H~M~2~1zinc~Fformalin_and_other_such_newer_fixatives~N_Can_a_zinc_salt~Kbr_~H~M~2~1really_replace_mercuric_chloride~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answers.~Kbr_~H~M~2~1~Kbr_~H~M~2~1These_questions_are_discussed_quite_frequently_in_the_HistoNet~Kbr_~H~M~2~1listserver_group._In_February_1998._I_wrote_that_there_was_a~Kbr_~H~M~2~1shortage_of_publications_in_refereed_journals~E_and_also~Kbr_~H~M~2~1suggested_that_it_was_unwise_to_use_a_commercial_product~Kbr_~H~M~2~1without_knowing_its_complete_composition._~AThere_are~Kbr_~H~M~2~1published_formulations~E_but_in_most_cases_these_compare_a~Kbr_~H~M~2~1zinc~Fcontaining_liquid_with_neutral_buffered_formaldehyde~E_for~Kbr_~H~M~2~1immunohistochemical_detection_of_one_or_several_antigens._The~Kbr_~H~M~2~1exact_composition_of_proprietary_fixative_mixtures_is_rarely~Kbr_~H~M~2~1stated_in_catalogues_etc.~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1John_Kiernan~Kbr_~H~M~2~1London~E_Canada~Kbr_~H~M~2~1~Akiernan~PAT~Quwo.ca~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1Dick_Dapson_disagreed_with_some_of_my_comments~E_and_provided~Kbr_~H~M~2~1a_helpful_list_of_publications~I~Kbr_~H~M~2~1~Kbr_~H~M~2~1John_Kiernan_wrote_~A2~H19~H98~B_that_there_is_a_remarkable_shortage~Kbr_~H~M~2~1of_literature_comparing_zinc_formalin_solutions_with_conventional~Kbr_~H~M~2~1fixatives._Actually~E_the_subject_has_been_covered_rather_well_over~Kbr_~H~M~2~1a_time_span_of_more_than_10_years._Here_is_a_sample_that_shows~Kbr_~H~M~2~1the_evolution_of_these_remarkable_fixatives~J_all_are_from_refereed~Kbr_~H~M~2~1journals_and_~Aexcept_for_the_1981_abstract~B_have_~4passed_the~Kbr_~H~M~2~1scrutiny_of_the_regular_scientific_publication_process~4~I~Kbr_~H~M~2~1~Kbr_~H~M~2~11981._Jones~E_et_al._Transition_metal_salts_as_adjuncts_to~Kbr_~H~M~2~1formalin_for_tissue_fixation_~Aabstract~B._Lab_Invest_44~I32A~Kbr_~H~M~2~1~PThis_is_the_paper_that_really_started_it_all~E_although_zinc~Kbr_~H~M~2~1formulations_do_appear_in_the_early_literature~Q.~Kbr_~H~M~2~1~Kbr_~H~M~2~11983._Mugnaini_et_al._Zinc~Faldehyde_fixation_for~Kbr_~H~M~2~1light~Fmicroscopic_immunocytochemistry_of_nervous_tissues.~Kbr_~H~M~2~1J_Histoch_Cytochem_31~I1435~F1438.~Kbr_~H~M~2~1~Kbr_~H~M~2~11985._Banks._Technical_aspects_of_specimen_preparation_and~Kbr_~H~M~2~1special_studies._In_Surgical_Pathology_of_the_Lymph_Nodes_and~Kbr_~H~M~2~1Related_Organs._Jaffe~E_ed._W_B_Saunders_Co.~E_pp1~F21.~Kbr_~H~M~2~1~Kbr_~H~M~2~11988._Herman~E_et_al._Zinc_formalin_fixative_for_automated~Kbr_~H~M~2~1tissue_processing._J_Histotechnol_11~I_85~F89.~Kbr_~H~M~2~1~PThe_first_really_comprehensive_study_comparing_NBF_and~Kbr_~H~M~2~1unbuffered_zinc_sulfate_formalin~Q.~Kbr_~H~M~2~1~Kbr_~H~M~2~11990._Tome~E_et_al._Preservation_of_cluster_1_small_cell_lung~Kbr_~H~M~2~1cancer_antigen_in_zinc~Fformalin_fixative_and_its_application_to~Kbr_~H~M~2~1immunohistochemical_diagnosis._Histopathol_16~I_469~F474.~Kbr_~H~M~2~1~Kbr_~H~M~2~11991._Abbondanzo~E_et_al._Enhancement_of_immunoreactivity_among~Kbr_~H~M~2~1lymphoid_malignant_neoplasms_in_paraffin~Fembedded_tissues_by~Kbr_~H~M~2~1refixation_in_zinc_sulfate~Fformalin._Arch_Pathol_Lab_med~Kbr_~H~M~2~1115~I31~F33.~Kbr_~H~M~2~1~Kbr_~H~M~2~11993._Estrogen_and_progesterone_receptor_proteins_in_zinc~Kbr_~H~M~2~1sulfate~E_formalin_fixed_breast_carcionoma~I_advantages_of_a~Kbr_~H~M~2~1supersensitive_streptavidin_technique._J_Histotechnol_16~I~Kbr_~H~M~2~151~F56.~Kbr_~H~M~2~1~Kbr_~H~M~2~11993._Dapson._Fixation_for_the_1990~9s~I_a_review_of_needs_and~Kbr_~H~M~2~1accomplishments._Biotechnic_~8amp~J_Histochem_68~I75~F82.~Kbr_~H~M~2~1~PLike_Herman~9s_paper~E_this_provides_a_critical_comparison~Kbr_~H~M~2~1between_NBF_and_zinc_formalin~J_it_also_details_probable~Kbr_~H~M~2~1mechanisms_and_reviews_the_pertinent_literature_to_date~Q.~Kbr_~H~M~2~1~Kbr_~H~M~2~11995._L~9Hoste~E_et_al._Using_zinc_formalin_as_a_routine~Kbr_~H~M~2~1fixative_in_the_histology_laboratory._Lab_Med_26~I_210~F214.~Kbr_~H~M~2~1~PCompares_NBF_and_a_buffered_zinc_formalin~E_using_side~Fby~Fside~Kbr_~H~M~2~1color_photomicrographs~Q.~Kbr_~H~M~2~1~Kbr_~H~M~2~1My_response~I~Kbr_~H~M~2~1~Kbr_~H~M~2~1The_interested_reader_should_study_these_publications._Most_do~Kbr_~H~M~2~1not_include_critical_comparisons_with_other_fixatives_~Aexcept~Kbr_~H~M~2~1buffered_formaldehyde~B~E_especially_for_preservation_of~Kbr_~H~M~2~1intracellular_structures._There_is_a_real_need_for_users_to~Kbr_~H~M~2~1compare_several_fixatives_in_properly_controlled_trials~E_and~Kbr_~H~M~2~1publish_their_results.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Zinc_mixtures_became_popular_in_the_early_1990s~E_but_the~Kbr_~H~M~2~1earliest_~Aprobably~B_of_its_kind_was_introduced_soon_after_the~Kbr_~H~M~2~1fixative_action_of_formaldehyde_was_discovered_by_F._Blum_~Ain~Kbr_~H~M~2~1Germany~E_in_1893~B._This_is_Fish~9s_fixative~I~Kbr_~H~M~2~1~Kbr_~H~M~2~1Water~I_2000_ml~Kbr_~H~M~2~1Formalin~I_50_ml~Kbr_~H~M~2~1Zinc_chloride~I_15_g~Kbr_~H~M~2~1~Kbr_~H~M~2~1Fish~E_Pierre_A._1895._The_use_of_formalin_in_neurology.~Kbr_~H~M~2~1Trans._Am._Microsc._Soc._17~I_319~F330.~Kbr_~H~M~2~1~PFish_recommended_immersion_of_the_brain_for_7~F10_days~E_with~Kbr_~H~M~2~1injection_of_cavities_and_blood_vessels_if_possible._It~9s_all~Kbr_~H~M~2~1been_done_before_if_you_go_back_far_enough~3_Fish~9s_paper_also~Kbr_~H~M~2~1reviewed_the_uses_of_formaldehyde_~A31_references~E_2_years~Kbr_~H~M~2~1after_it~9s_introduction_as_a_fixative~B_and_described_other~Kbr_~H~M~2~1fixative_mixtures.~Q~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Alternatives_to_mercury~Fcontaining_fixatives~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1What_is_the_best_substitute_for_B~F5_fixative~E_without~Kbr_~H~M~2~1mercuric_chloride~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1~P_B~F5_is~I_Water_90_ml~Kbr_~H~M~2~1Formalin_~A40~7_HCHO~B_10_ml~Kbr_~H~M~2~1Mercuric_chloride_6_g~Kbr_~H~M~2~1Sodium_acetate_~Aanhydrous~B_1.25_g~Kbr_~H~M~2~1The_sodium_acetate_brings_the_pH_into_the_5.8~F6.0_range.~Kbr_~H~M~2~1Fix_by_immersion~E_12~F24_hours~E_then_transfer_to_70~F80~7~Kbr_~H~M~2~1alcohol._See_Lillie_RD_~8amp~J_Fullmer_HM_1976_Histopathologic~Kbr_~H~M~2~1Technic_and_Practical_Histochemistry._New_York~I~Kbr_~H~M~2~1McGraw~FHill~E_pp_52~F53.~Q~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1We_recently_completed_a_~4blind_comparison~4_of_B~F5_substitutes._We_needed~Kbr_~H~M~2~1to_find_something~E_as_our_water_treatment_plant_had_notified_us_that_as~Kbr_~H~M~2~1part_of_a_Zero_Discharge_Program_they_would_be_monitoring_our_mercury~Kbr_~H~M~2~1output._Of_course~E_we_were_capturing_our_mercury_..._but_we_still_had~Kbr_~H~M~2~1measurable_amounts_in_our_discharged_water._The_treatment_plant~Kbr_~H~M~2~1immediately_zeroed_in_on_our_department~E_and_without_delay_asked_if_we~Kbr_~H~M~2~1used_mercury_fixatives~3_We_agreed_that_we_would_cease~E_or_absolutely~Kbr_~H~M~2~1contain_our_mercury_by_June_1~E_1998._I_felt_it_better_to_cease_using~Kbr_~H~M~2~1mercury~E_so_that_any_future_mercury_found_in_the_discharge_water_from~Kbr_~H~M~2~1the_hospital_could_be_blamed_on_another_source~3~3_We_had_all_of_our_sink~Kbr_~H~M~2~1traps_cleaned~E_and_tested_..._no_mercury_coming_from_us~3~3~3~Kbr_~H~M~2~1~Kbr_~H~M~2~1For_our_study~E_we_used_our_standard_B~F5~E_Z~F5~E_and_Z~Ffix_from_Anatech~E~Kbr_~H~M~2~1IBF_from_Surgipath~E_our_10~7_NBF~E_and_B~Fplus_fixative_from_***._We_used~Kbr_~H~M~2~1tonsil_and_lymph_nodes_for_the_study~E_and_placed_small_pieces_of_tissue~Kbr_~H~M~2~1in_each_of_the_fixatives~E_and_gave_them_to_the_pathologists_labeled_as~Kbr_~H~M~2~1fixative_1~E_fixative_2_etc._The_pathologists_were_given_an_evaluation~Kbr_~H~M~2~1sheet_with_each_case~E_and_asked_to_rank_the_fixatives_from_1~F6~E_with_1~Kbr_~H~M~2~1being_the_best._When_we_had_tested_a_sufficient_number_of_cases~E_the~Kbr_~H~M~2~1evaluations_were_tallied~E_and_lo_and_behold_..._B~F5_won~3_I_wasn~9t~Kbr_~H~M~2~1surprised~E_and_neither_were_the_pathologists._We_all_agreed_that_we~Kbr_~H~M~2~1would_use_the_second_place_winner.~Kbr_~H~M~2~1~Kbr_~H~M~2~1This_was_B~FPlus_Fix_which_is_sold_by_***_~A800~F635~F4477~E_or_write_to_PO~Kbr_~H~M~2~1Box_609~E_Stanwood~E_WA_98292~B._However~E_all_the_solutions_that_we_tested~Kbr_~H~M~2~1were_acceptable._One_surprising_result_was_that_our_10~7_NBF_came_in_3rd~E~Kbr_~H~M~2~1very_close_to_our_2nd_place_winner._We_have_been_using_our_substitute~Kbr_~H~M~2~1since_March~E_and_are_pleased_with_the_results_so_far..._However~E_the~Kbr_~H~M~2~1pathologists_are_missing_their_B~F5~E_which_they_still_refer_to_as_the~Kbr_~H~M~2~1gold_standard.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~F~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1PROCESSING~E_DECALCIFYING~E_EMBEDDING~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Solvent_to_replace_xylene_AND_alcohols~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Is_there_a_product_that_replaces_xylene_AND_alcohols_in_the~Kbr_~H~M~2~1staining_procedure~N_Can_you_use_it_before_and_after_the_actual~Kbr_~H~M~2~1staining_is_done~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_1.~Kbr_~H~M~2~1~Kbr_~H~M~2~1t~Fbutanol~E_dioxane_and_tetrahydrofuran_are_miscible_with~Kbr_~H~M~2~1wax~E_water_and_resinous_mounting_media._Of_these~E_only~Kbr_~H~M~2~1t~Fbutanol_~A~L_tertiary_butyl_alcohol~B_is_suitable_for~Kbr_~H~M~2~1ordinary_use._~AThe_other_two_have_such_hazards_as_fire~E~Kbr_~H~M~2~1toxicity_and_explosive_peroxide_formation.~B_t~Fbutanol_is~Kbr_~H~M~2~1often_used_in_botanical_microtechnique~J_it_is_quite_a_bit~Kbr_~H~M~2~1more_expensive_than_alcohol_or_xylene._n~Fbutyl_alcohol~Kbr_~H~M~2~1mixes_with_wax_and_mounting_media_and_is_also_partly~Kbr_~H~M~2~1miscible_with_water._It~9s_good_when_you_use_easily~Kbr_~H~M~2~1extracted_stains_~Amethyl_green~Fpyronine~E_for_example~B~E~Kbr_~H~M~2~1but_has_unpleasant_vapour.~Kbr_~H~M~2~12~Fbutoxyethanol_~Abutyl_cellosolve~B_also_has_the_right~Kbr_~H~M~2~1miscibilities~E_and_is_quite_cheap_because_it~9s_used~Kbr_~H~M~2~1on_a_big_scale_industrially.~Kbr_~H~M~2~1For_microwave_processing~E_isopropyl_alcohol_is~Kbr_~H~M~2~1sometimes_recommended._However~E_this_does_not_mix~Kbr_~H~M~2~1with_wax._It_has_to_leave_the_specimen_by_vaporizing~Kbr_~H~M~2~1~Aboiling~B_under_reduced_pressure._This_can_lead_to~Kbr_~H~M~2~1considerable_tissue_damage_unless_the_temperature~Kbr_~H~M~2~1and_pressure_are_just_right_~ABosch_et_al_1996~B.~Kbr_~H~M~2~1Some_staining_methods_work_well~E_though_slowly~E_without~Kbr_~H~M~2~1removing_the_paraffin_beforehand_~AKiernan_1996~B~E_provided~Kbr_~H~M~2~1that_there_has_been_no_melting_or_softening_of_the_wax~Kbr_~H~M~2~1after_mounting_the_sections_on_their_slides.~Kbr_~H~M~2~1~Kbr_~H~M~2~1References.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Bosch~EMMC~J_Walspaap~ECH~J_Boon~EME_~A1996~B~I_Lessons_from_the~Kbr_~H~M~2~1experimental_stage_of_the_two~Fstep_vacuum~Fmicrowave_method~Kbr_~H~M~2~1for_histoprocessing._Eur._J._Morphol._34~A2~B~E_127~F130.~Kbr_~H~M~2~1Kiernan~EJA_~A1996~B~I_Staining_paraffin_sections_without_prior~Kbr_~H~M~2~1removal_of_the_wax._Biotechnic_~8amp~J_Histochemistry_71~A6~B~E~Kbr_~H~M~2~1304~F310.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_2.~Kbr_~H~M~2~1~Kbr_~H~M~2~1We_use_99~7_isopropyl_alcohol_~AIPA~B_instead_ethanol_AND_xylene~Kbr_~H~M~2~1AFTER_staining._It_is_especially_useful_after_staining_of_lymph~Kbr_~H~M~2~1nodes_with_a_modified_Maximov~FGiemsa_method._My_laboratory_has~Kbr_~H~M~2~1used_this_modification_more_then_5_years_and_I_have_never_seen~Kbr_~H~M~2~1the_same_excellent_result_in_comparison_with_atlases_of_lymph~Kbr_~H~M~2~1nodes_biopsy._Moreover~E_we_use_IPA_with_addition_of_a_small~Kbr_~H~M~2~1amount_of_detergent_for_dehydration_of_samples._Four_changes_of~Kbr_~H~M~2~199~7_IPA~Ddetergent_is_all_you_need_between_water_and_paraffin.~Kbr_~H~M~2~1We_never_have_have_problems_with_any_tissues~E_including_large~Kbr_~H~M~2~1samples_of_skin._Our_HTs_adore_IPA.~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_2~Fbutoxyethanol_~A~4Clereum~4~B_dehydrating_or_clearing_agent~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1What_are_the_properties_of_Clereum~N_~AThe_MSDS_for_Clereum~Kbr_~H~M~2~1indicates_the_ingredient_information_as_undiluted~Kbr_~H~M~2~12~Fbutoxyethanol.~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1It~9s_good_to_learn_that_this_isn~9t_yet_another_secret_clearing~Kbr_~H~M~2~1agent~3_According_to_the_Merck_Index~E_this_compound_~Aalso_called~Kbr_~H~M~2~1butyl_cellosolve~E_or_ethylene_glycol_monobutyl_ether~B_is_partly~Kbr_~H~M~2~1miscible_with_water._Its_properties_as_a_solvent_seem_to_be~Kbr_~H~M~2~1similar_to_n~Fbutanol~J_no_doubt_the_higher_B.P._~A171C~B_is_an~Kbr_~H~M~2~1advantage_~F_it_won~9t_have_n~Fbutanol~9s_nasty_cough~Fmaking_vapour.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Merck_says_the_toxicity_is_similar_to_methyl_cellosolve~Kbr_~H~M~2~1~Aanaemia~E_~4CNS_symptoms~4_etc~J_can_be_absorbed_through_skin~B.~Kbr_~H~M~2~1~Kbr_~H~M~2~1The_price_of_2~Fbutoxyethanol_varies_with_the_supplier_~AMay_1997~B~I~Kbr_~H~M~2~1~Kbr_~H~M~2~1Fisher_Scientific_4_litres_$104_~A~4Laboratory_grade~4~B~Kbr_~H~M~2~1Sigma_3_kg_$42_~Ano_purity_details~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1Acros_Organics_~Aseems_to_be_part_of_Fisher~B~Kbr_~H~M~2~1sell_three_grades~I~Kbr_~H~M~2~12.5_litres_$24_~A99~7~B~Kbr_~H~M~2~11_kg_$23_~AGC~B~Kbr_~H~M~2~1500_ml_$36_~Ascintillation_grade~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1If_the_99~7_stuff_is_OK_for_histology~E_perhaps_the_price_isn~9t~Kbr_~H~M~2~1too_bad._tert~Fbutanol_~A99.5~7~J_from_Acros~B_is_$67_for_2.5_litres~E~Kbr_~H~M~2~1and_n~Fbutanol_~A99~7~B_is_$27_for_2.5_litres._This_makes~Kbr_~H~M~2~12~Fbutoxyethanol_quite_a_good_buy_for_a_non~Fniffy~Kbr_~H~M~2~1not~Fquite~Funiversal_solvent._The_similarity_of_its_miscibilities~Kbr_~H~M~2~1to_those_of_n~Fbutanol_suggests_that_this_might_be_useful_for~Kbr_~H~M~2~1dehydrating_~Aand_clearing~B_sections_that_have_been_stained_with~Kbr_~H~M~2~1methyl_green~Fpyronine~E_or_other_dyes_that_are_easily_lost_with~Kbr_~H~M~2~1ordinary_alcoholic_dehydration.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Decalcification~I_Acid_or_EDTA~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Questions.~Kbr_~H~M~2~1~Kbr_~H~M~2~1How_should_I_decalcify_a_bony_specimen_or_a_tooth~N~Kbr_~H~M~2~1What_precautions_are_needed_if_galactosidase_activity_must_be~Kbr_~H~M~2~1preserved_~Ato_identify_cells_carrying_the_LacZ_gene~B~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_1.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Decalcification_with_EDTA_is_probably_the_best_method_with_your~Kbr_~H~M~2~1LacZ~E_due_to_the_enzyme_staining_you_are_doing._I_would_be~Kbr_~H~M~2~1careful_to_adjust_the_pH_of_the_EDTA_solution_to_the_working_pH~Kbr_~H~M~2~1of_enzyme_staining_in_PBS_or_a_TRIS_buffer~E_and_rinse_carefully~Kbr_~H~M~2~1in_buffers_postdecalcification._Formic_acid_may_ruin_LacZ~Kbr_~H~M~2~1enzyme_staining_results.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Gayle_Callis~Kbr_~H~M~2~1~Auvsgc~PAT~Qmsu.oscs.montana.edu~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_2.~Kbr_~H~M~2~1~Kbr_~H~M~2~1If_the_bone_is_crunchy~E_you_have_either_not_removed_all_the_bone~Kbr_~H~M~2~1mineral~E_or_you_have_transferred_the_bones_from_EDTA_to_alcohol~Kbr_~H~M~2~1and_have_precipitated_EDTates_in_your_tissue.~Kbr_~H~M~2~1~Kbr_~H~M~2~1When_you_decalcify~E_do_you_determine_the_end_point_using_an_x~F~Kbr_~H~M~2~1ray~Hcalcium_oxalate~Hprod_with_a_pointed_stick~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1How_long_do_you_decalcify~N_Even_at_20~7_EDTA_these_would_take_at~Kbr_~H~M~2~1least_a_week_with_vigorous_agitation_at_room_temperature._Is~Kbr_~H~M~2~1the_EDTA_buffered_to_pH_7~N_If_not~E_you_are_using_the_solution~Kbr_~H~M~2~1as_an_acid_decalcifier_as_well_as_a_chelator._In_this_case~E~Kbr_~H~M~2~1assuming_your_stain_still_works_and_will_not_be_affected_by_acid~Kbr_~H~M~2~1pH~E_change_to_10~7_formic_acid~E_which_provides_much_faster~Kbr_~H~M~2~1decalcification._Check_the_endpoint_~Awhen_all_the_calcium_is~Kbr_~H~M~2~1gone~B_daily.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~P_But_see_Answer_1_for_acid~Fsensitivity_of_galactosidase._~Q~Kbr_~H~M~2~1~Kbr_~H~M~2~1If_you_have_checked_the_endpoint_and_all_the_calcium_is_gone~E~Kbr_~H~M~2~1rinse_the_tissue_in_water_for_at_least_8_hours_to_remove_all_the~Kbr_~H~M~2~1excess_EDTA_before_putting_it_in_alcohol.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_3._~AA_formic_acid_procedure_for_teeth~E_with_oxalate_testing~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1The_protocol_we_use_here_at_Ind._Univ._School_of_Dentistry_is_as~Kbr_~H~M~2~1follows~I~Kbr_~H~M~2~1~Kbr_~H~M~2~1The_protocol_we_use_here_at_Ind._Univ._School_of_Dentistry_is_as~Kbr_~H~M~2~1follows~I~Kbr_~H~M~2~1~Kbr_~H~M~2~1After_teeth_are_fixed_in_10~7_neutral_buffered_formalin~E_they_are~Kbr_~H~M~2~1placed_in_wide_mouth_bottles_with_a_5~7_formic_Acid_solution.~Kbr_~H~M~2~1They_are_then_checked_each_day_by_pipetting_5_ml_of_the_acid~Kbr_~H~M~2~1solution_into_a_test_tube_to_which_1_ml_of_2.5~7_ammonium_oxalate~Kbr_~H~M~2~1is_added._If_a_white_precipitate_forms_there_is_still_calcium~Kbr_~H~M~2~1present._The_solution_is_then_changed_and_the_process_repeated~Kbr_~H~M~2~1the_next_day._Once_I_get_one_negative_test_the_specimen_is~Kbr_~H~M~2~1grossed_as_needed_and_placed_back_into_acid_until_another~Kbr_~H~M~2~1negative_is_obtained._The_specimen_is_then_placed_in_running~Kbr_~H~M~2~1water_overnight_and_processed_with_the_next_days_run._I_know~Kbr_~H~M~2~1this_can_take_a_long_time~E_but_the_results_are_worth_it._If_you~Kbr_~H~M~2~1need_anything_else_let_me_know.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Testing_for_completeness_of_decalcification~Kbr_~H~M~2~1~Kbr_~H~M~2~1Questions.~Kbr_~H~M~2~1~Kbr_~H~M~2~1How_should_I_test_for_complete_decalcification~N~Kbr_~H~M~2~1Is_the_same_method_OK_after_either_formic_acid~Kbr_~H~M~2~1or_EDTA~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_2.~Kbr_~H~M~2~1~Kbr_~H~M~2~1The_ammonium_oxalate_test_is_simple._Take_a_5_ml_sample~Kbr_~H~M~2~1of_used_decalcifying_fluid._Neutralize_it_by_adding_drops~Kbr_~H~M~2~1of_strong_ammonia_~Aammonium_hydroxide~B~J_avoid_the_fumes~3~Kbr_~H~M~2~1When_the_solution_turns_litmus_blue_~ApH_above_7~B~E~Kbr_~H~M~2~1add_5_ml_of_saturated_aqueous_solution_of_ammonium~Kbr_~H~M~2~1oxalate_~Aabout_3~7~J_stable_stock_solution~B._Wait_for~Kbr_~H~M~2~1half_an_hour._If_there_is_no_precipitate~E_the_last~Kbr_~H~M~2~1change_of_decalcifying_fluid_was_free_of_calcium_ions.~Kbr_~H~M~2~1~Kbr_~H~M~2~1According_to_Eggert_~8amp~J_Germain_~A1979~B_you_can_use_the_ammonium~Kbr_~H~M~2~1oxalate_test_on_EDTA._Rosen_~A1981~B_said_the_sensitivity_was~Kbr_~H~M~2~1higher_if_you_lowered_the_pH_to_3.2~F3.6_before_doing_the_test~Kbr_~H~M~2~1~Ainstead_of_neutralizing_to_pH_7_as_done_with_an_acid_decalcifier~B.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Eggert_FM~E_Germain_JP_1979._Histochemistry_39~I_215~F224.~Kbr_~H~M~2~1Rosen_AD_1981._End~Fpoint_determination_in_EDTA_decalcification~Kbr_~H~M~2~1using_ammonium_oxalate._Stain_Technology_56~I_48~F49.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Fatty_specimens~I_Processing_into_paraffin.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1What_is_the_best_way_to_paraffin~Fembed_specimens_that~Kbr_~H~M~2~1contain_a_lot_of_fat~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answers.~Kbr_~H~M~2~1~Kbr_~H~M~2~11._Process_by_hand~E_allowing_more_time_and_bigger_volumes~Kbr_~H~M~2~1of_all_solvents_than_for_non~Ffatty_pieces_of_tissue.~Kbr_~H~M~2~12._Don~9t_put_them_through_an_automatic_processor_because~Kbr_~H~M~2~1you~9ll_get_grease_in_all_the_solvents._~AIf_you_don~9t~Kbr_~H~M~2~1believe_this~E_put_a_bit_of_skin_in_about_10_times_its~Kbr_~H~M~2~1volume_of_95~7_alcohol_for_an_hour~E_then_add_some~Kbr_~H~M~2~1water_to_the_alcohol._Result~I_a_milky_emulsion.~B~Kbr_~H~M~2~13._Xylene_is_better_than_a_~4xylene_substitute.~4~Kbr_~H~M~2~1~Kbr_~H~M~2~1~P_Unfortunately_I_mislaid_the_sources_of_these_pieces_of~Kbr_~H~M~2~1advice._For_what_it~9s_worth~E_I_agree_strongly_with~Kbr_~H~M~2~1the_first_two~E_but_lack_the_experience_to_comment_on~Kbr_~H~M~2~1the_third._J._A._Kiernan._~Q~Kbr_~H~M~2~1~Kbr_~H~M~2~1Back_to_Table_of_Contents~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Polymethyl_methacrylate_embedding_for_bone~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Is_it_permissible_to_mix_polymerized_methyl_methacrylate~Kbr_~H~M~2~1with_the_monomer~E_when_making_an_embedding_medium_for~Kbr_~H~M~2~1undecalcified_bone~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Using_polymethylmethacrylate_powder_or_beads_does_not_affect~Kbr_~H~M~2~1the_polymerization_process~E_but_it_does_make_the_preparation~Kbr_~H~M~2~1of_the_partly_polymerized_embedding_mixture_easier_and_safer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1You_may_care_to_refer_to_the_following_paper.~Kbr_~H~M~2~1Difford~E_J._~A1974~B_~4A_simplified_method_for_the_preparation~Kbr_~H~M~2~1of_methyl_methacrylate_embedding_medium_for_undecalcified~Kbr_~H~M~2~1bone.~4_Medical_Laboratory_Technology_31~I_79~F81.~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Mold_release_spray~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Is_there_something_you_can_spray_into_an_embedding~Kbr_~H~M~2~1mold_to_make_it_easier_to_extract_the_solidified~Kbr_~H~M~2~1wax_block~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1I_faced_the_problem_of_mold~Frelease_spray_several_years_ago~Kbr_~H~M~2~1by_mixing_a_solution_of_5~7_green_dishwashing_soap_~Asuch_as~Kbr_~H~M~2~1Palmolive~B_in_50~7_Ethanol~E_then_putting_it_into_a_pump_spray~Kbr_~H~M~2~1bottle_~Aavailable_form_any_housewares_department~B._This~Kbr_~H~M~2~1worked_AT_LEAST_as_well_as_the_outrageously_expensive_stuff~Kbr_~H~M~2~1sold_as_~4Mold~FRelease_Spray~4~E_and_it_contained_no_CFC~9s_or~Kbr_~H~M~2~1other_~4evils~4.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Paraffin_processing_of_skin~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Could_you_suggest_a_processing_schedule_suitable_for_skin~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1This_is_my_processing_schedule_for_skin_dehydration_and~Kbr_~H~M~2~1embedding.~Kbr_~H~M~2~1~Kbr_~H~M~2~1By_Hand.~Kbr_~H~M~2~1~Kbr_~H~M~2~1The_times_suit_my_working_day._I~9m_sure_they_could_be~Kbr_~H~M~2~1altered_for_any_work_pattern.~Kbr_~H~M~2~1~Kbr_~H~M~2~11.~B_80~7_alcohol._~L_2_pm.~Kbr_~H~M~2~12.~B_80~7_alcohol._~L_5_pm_~F_overnight.~Kbr_~H~M~2~13.~B_Abs.alc.~H8~7_phenol._~L_9_am.~Kbr_~H~M~2~14.~B_Abs._alcohol._~L_10_am.~Kbr_~H~M~2~15.~B_Abs._alcohol._~L_12_am.~Kbr_~H~M~2~16.~B_Abs.alc.~Hamyl_acetate.~L_3_pm.~Kbr_~H~M~2~17.~B_Amyl_acetate._~L_4_pm.~Kbr_~H~M~2~18.~B_Amyl_acetate._~L_5_pm._~F_overnight.~Kbr_~H~M~2~19.~B_Amyl_acetate._~L_9_am.~Kbr_~H~M~2~110.~B_Amyl_acetate._~L_12_am.~Kbr_~H~M~2~111.~B_Xylene._~L_3_pm.~Kbr_~H~M~2~112.~B_Wax._~L_4_pm.~Kbr_~H~M~2~113.~B_Wax._~L_5pm._~F_overnight.~Kbr_~H~M~2~114.~B_Wax._~L_Embed.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Tissue_Processor.~Kbr_~H~M~2~1~Kbr_~H~M~2~1These_times_I_use_on_a_Shandon_Histokinette~E_remember~Kbr_~H~M~2~1them_~N.~Kbr_~H~M~2~1~Kbr_~H~M~2~11.~B_80~7_alcohol._~L_2_hours.~Kbr_~H~M~2~12.~B_80~7_alcohol._~L_2_hours.~Kbr_~H~M~2~13.~B_Abs._alc.~H8~7_phenol._~L_1_hour.~Kbr_~H~M~2~14.~B_Abs._alcohol._~L_3_hours.~Kbr_~H~M~2~15.~B_Abs._alcohol._~L_3_hours.~Kbr_~H~M~2~16.~B_Abs.alc.~Hamyl_acetate.~L_1_hour.~Kbr_~H~M~2~17.~B_Amyl_acetate._~L_3_hours.~Kbr_~H~M~2~18.~B_amyl_acetate._~L_3_hours.~Kbr_~H~M~2~19.~B_Amyl_acetate._~L_3_hours.~Kbr_~H~M~2~110.~B_Amyl_acetate._~L_5_hours.~Kbr_~H~M~2~111.~B_Xylene._~L_1_hour.~Kbr_~H~M~2~112.~B_Wax._~L_9_hours.~Kbr_~H~M~2~113.~B_Wax._~L_9_hours.~Kbr_~H~M~2~114.~B_Embed.~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Cryoprotection_of_specimens~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Please_recommend_a_way_to_protect_formaldehyde~Ffixed~Kbr_~H~M~2~1mouse_brains_to_avoid_crack_and_ice_crystal_holes~Kbr_~H~M~2~1that_for_during_rapid_freezing._25~7_sucrose_has_been~Kbr_~H~M~2~1recommended._Should_it_be_in_water_or_phosphate~Kbr_~H~M~2~1buffered_saline~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_1.~Kbr_~H~M~2~1~Kbr_~H~M~2~1For_ultracryomicrotomy_~Aor_should_it_be_cryoultramicrotomy~B~Kbr_~H~M~2~1Tokuyasu_~A1989~B_used_2.3_M_~A~L_78~7~B_sucrose_in_0.1M_phosphate_buffer.~Kbr_~H~M~2~1He_was_working_with_blocks_much_smaller_than_mouse_brain~E_so_you~Kbr_~H~M~2~1will_no_doubt_have_to_increase_the_time._Inflitration_of_blocks~Kbr_~H~M~2~11_mm_wide_usually_took_30_minutes._He_stated_that_infusion_was~Kbr_~H~M~2~1complete_when_the_specimen_no_longer_floated_on_the_top_of~Kbr_~H~M~2~1the_sucrose_solution._The_same_author_reported_that~Kbr_~H~M~2~110~F30~7_PVP_and_1.6~F2M_sucrose_provided_still_better_postfreezing~Kbr_~H~M~2~1conditions_~Acompared_with_freezing_alone~B.~Kbr_~H~M~2~1~Kbr_~H~M~2~1We_presently_use_5~7_PVA_~Apolyvinyl_alcohol~B_in_phosphate_buffer~Kbr_~H~M~2~1to_cryoprotect_bone_samples_before_freezing_for_enzyme_and~Kbr_~H~M~2~1immunohistochemistry.~Kbr_~H~M~2~1~Kbr_~H~M~2~1One_other_point_that_may_be_worth_considering_is_the_method~Kbr_~H~M~2~1for_freezing._If_you_are_thinking_of_snap~Ffreezing~E_I_would~Kbr_~H~M~2~1recommend_hexane_instead_of_isopentane._Hexane_freezes_at_a~Kbr_~H~M~2~1considerably_higher_temperature~I_about_80_C._Many_moons_ago~E~Kbr_~H~M~2~1when_I_worked_in_Neuropathology_in_Scotland~E_I_found_that_mouse~Kbr_~H~M~2~1brains_tended_to_crack_when_frozen_in_isopentane~E_but_that_we~Kbr_~H~M~2~1had_much_better_preservation_when_freezing_in_precooled_hexane~Kbr_~H~M~2~1~Awe_never_cryoprotected_them_though~B.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_2.~Kbr_~H~M~2~1~Kbr_~H~M~2~1It_is_a_common_practice_to_immerse_rodent_brains_in_20~F30~7_sucrose~Kbr_~H~M~2~1at_4_C~E_at_least_until_they_sink._If_they_have_been_fixed_for~Kbr_~H~M~2~1only_a_short_time_~Aless_than_48_hours~B~E_it_is_probably_best_to~Kbr_~H~M~2~1dissolve_the_sucrose_in_PBS_rather_than_water_alone.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Rosene_et_al_~A1986~B_found_that_20~7_glycerol_with~Kbr_~H~M~2~12~7_dimethylsulfoxide_~ADMSO~B_was_better_than_sucrose.~Kbr_~H~M~2~1The_sucrose_concentration_needs_to_be_much_higher_than~Kbr_~H~M~2~1is_commonly_used_~F_at_leased_60~7_~Asee_Lepault_et_al~E~Kbr_~H~M~2~11997~B.~Kbr_~H~M~2~1~Kbr_~H~M~2~1References_~Awith_brief_notes~B.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Rosene~EDL~J_Roy~ENJ~J_Davis~EBJ_~A1986~B~I_A_cryoprotection_method~Kbr_~H~M~2~1that_facilitates_cutting_frozen_sections_of_whole_monkey~Kbr_~H~M~2~1brains_for_histological_and_histochemical_processing~Kbr_~H~M~2~1without_freezing_artifact._J._Histochem._Cytochem._34~E~Kbr_~H~M~2~11301~F1315.~Kbr_~H~M~2~1Techniques_compared._Optimum_cryoprotection_with_4_day~Kbr_~H~M~2~1infiltration_~A4_C~B_of_20~7_glycerol_~8amp~J_2~7_DMSO_in_buffer~Kbr_~H~M~2~1or_fixative._Then_freeze_in_isopentane_at_~F75_C_~Adry~Kbr_~H~M~2~1ice~B._Better_than_other_cryoprotectants_~Asucrose_etc~B~Kbr_~H~M~2~1and_freezing_methods.~Kbr_~H~M~2~1Lepault~EJ~J_Bigot~ED~J_Studer~ED~J_Erk~EI_~A1997~B~I_Freezing_of_aqueous~Kbr_~H~M~2~1specimens~I_an_X~Fray_diffraction_study._J._Microsc.~Kbr_~H~M~2~1~AOxford~B_187~ASep~B~E_158~F166.~Kbr_~H~M~2~1EM_~8amp~J_X~Fray_diffraction_of_freezing_of_sucrose~Kbr_~H~M~2~1solutions._Immersion_in_a_liquid_cryogen_or_high~Kbr_~H~M~2~1pressure_freezing._Sucrose_favours_formation_of~Kbr_~H~M~2~1amorphous_ice~J_conc_must_be_60~7_or_above_for~Kbr_~H~M~2~1freezing_in_a_cryogenic_liquid.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Cutting_sections_of_toe_or_finger_nails~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Does_anyone_have_a_few_hints_for_sectoning_toenails~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1~P_Here_is_a_selection_of_many_replies_to_this~Kbr_~H~M~2~1frequently_asked_question._~Q~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_1.~Kbr_~H~M~2~1~Kbr_~H~M~2~110~7_Potassium_hydroxide._Soak_them_for_at_least~Kbr_~H~M~2~14_hrs~E_but_not_more_than_8.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Noreen_S._Gilman_~An4xiu~PAT~Qgate.net~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_2.~Kbr_~H~M~2~1~Kbr_~H~M~2~1I_have_not_cut_toenails_for_years._~AI_do_cut_my_own_personal~Kbr_~H~M~2~1toenails_of_course~3~B_However~E_we_used_to_soak_them_for_a_short~Kbr_~H~M~2~1time_in_Nair~E_which_i_believe_is_like_Neet~E_and_we_got_an~Kbr_~H~M~2~1excellent_section._~PSee_also_Answers_4_and_5.~Q~Kbr_~H~M~2~1~Kbr_~H~M~2~1The_procedure_is_to_process_the_nails~E_and_after_they_are~Kbr_~H~M~2~1embedded_treat_the_paraffin_block_by_putting_it_in_a_petri_dish~Kbr_~H~M~2~1containing_the_Nair._The_Nair_is_put_in_first_and_then_the_block~Kbr_~H~M~2~1is_put_on_top._We_treat_the_block_for_5~F10_minutes_depending_on~Kbr_~H~M~2~1the_size_of_the_nails._We_wipe_off_the_block~E_try_cutting_it_and~Kbr_~H~M~2~1put_it_back_for_further_treatment_if_needed._It_is_best_to_cool~Kbr_~H~M~2~1the_block_on_iced_water_after_treatment_and_before_cutting_and~Kbr_~H~M~2~1to_take_the_first_sections.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Marjorie_Hagerty~Kbr_~H~M~2~1~Amhagerty~PAT~Qemc.org~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_3.~Kbr_~H~M~2~1~Kbr_~H~M~2~1I_learned_a_new_technique_at_one_of_the_outstanding_workshops_at~Kbr_~H~M~2~1NSH~FAlbuquerque._Our_hospital_switched_to_this_method._After~Kbr_~H~M~2~1grossing~E_place_a_representative_piece_~Aor_ALL_if_melanoma_is~Kbr_~H~M~2~1indicated~B_in_a_cassette_and_immerse_the_nail_in_5~7_Tween_80~Kbr_~H~M~2~1~ASigma_cat~5P~F4634~B_for_1~F2_hours_at_least._Overnight_won~9t_hurt~Kbr_~H~M~2~1it._Then_remove_and_process_as_usual._I_find_that_if_you_orient~Kbr_~H~M~2~1the_nail_to_cut_it_perpendicular_to_the_knife_it_cuts_more~Kbr_~H~M~2~1easily._Use_a_charged_or_polylysine_slide_~Aor_Elmers_glue_if~Kbr_~H~M~2~1it~9s_really_likely_that_it_will_float~B.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_4.~Kbr_~H~M~2~1~Kbr_~H~M~2~1There_are_several_methods_in_Luna~9s_last_book_~4Histopathologic~Kbr_~H~M~2~1Methods_and_Color_Atlas_of_Special_Stains_and_Tissue_Artifacts~4~Kbr_~H~M~2~1for_softening_keratin_in_nails~E_etc._Fixation_in_10~7_buffered~Kbr_~H~M~2~1formalin_is_necessary_to_produce_crosslinking_and_thereby~Kbr_~H~M~2~1prevent_keratin_from_dissolving_completely_in_softening~Kbr_~H~M~2~1solutions._After_fixation_and_BEFORE_processing_~F~F_place~Kbr_~H~M~2~1specimen_in_~4Neet~4_or_other_depilatory_cream_or_permanent_wave~Kbr_~H~M~2~1solution_for_one_to_several_hours._The_key_ingredient_in_these~Kbr_~H~M~2~1solutions_is_thioglycollate._*_This_is_best_performed_under_a~Kbr_~H~M~2~1hood_because_these_products_smell_really_bad_and_will_guarantee~Kbr_~H~M~2~1an_increase_in_lab_traffic_by_interested_personnel_wanting_to~Kbr_~H~M~2~1know_~4What_on_earth_are_you_doing~N~4_The_specimen_should_bend~Kbr_~H~M~2~1easily_before_continuing_with_next_step._Wash_the_specimen_in~Kbr_~H~M~2~1running_tap_water_for_10_minutes._Dehydrate~E_clear~E_and~Kbr_~H~M~2~1impregnate_with_paraffin_as_desired._Processing_times_will~Kbr_~H~M~2~1depend_on_which_hoof_you_are_processing_~F~F_elephants_take_a_lot~Kbr_~H~M~2~1longer_than_goats_~I~F~B_Get_out_your_nose_clip_and_have_fun~3~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer_5.~Kbr_~H~M~2~1~Kbr_~H~M~2~1We_have_routinely_used_~4Neet~4_overnight_and_had_good_results.~Kbr_~H~M~2~1Recently_tried_~4Neet~4_at_58_C_~Ait_liquefies~B_for_several_hours~Kbr_~H~M~2~1during_the_day_on_a_particularly_tough_nail~J_it_cut_beautifully~Kbr_~H~M~2~1the_next_day~3~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Paraffin_wax~I_crystals~E_additives_and_cutting~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1What_are_the_best_polymers_or_other_additives_for~Kbr_~H~M~2~1reducing_crystal_size_and_improving_the_cutting~Kbr_~H~M~2~1propereties_of_paraffin_wax~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Paraffin_wax_is_a_mixture_of_~Avirtually~B_straight_chain~Kbr_~H~M~2~1hydrocarbons._Note_the_word_~4mixture~4._Unless_you_go_to~Kbr_~H~M~2~1enormous_lengths_~Aof_purifying_or_searching_for_a_fine_chemical~Kbr_~H~M~2~1supplier~B~E_you_will_ALWAYS_have_a_mixture._There_is_a~Kbr_~H~M~2~1relationship_between_hydrocarbon_chain_length_and_melting~Kbr_~H~M~2~1point~E_but_as_the_waxes_are_always_mixtures~E_melting_points_are~Kbr_~H~M~2~1never_exact~E_either_in_the_compounding_or_the_measuring~E_but~Kbr_~H~M~2~1that_is_another_story~3~Kbr_~H~M~2~1~Kbr_~H~M~2~1Perhaps_more_important_than_the_melting_point_is_the_~4plastic~Kbr_~H~M~2~1point~E~4_but_that_is_virtually_ignored_by_our_suppliers._The~Kbr_~H~M~2~1plastic_point_occurs_about_10_C_below_the_melting_point_and~Kbr_~H~M~2~1its_meaning_should_be_fairly_obvious_~F_try_softening_a_piece~Kbr_~H~M~2~1of_physiotherapy_wax_in_your_hands_and_that_should_explain_all~Kbr_~H~M~2~1you_need_to_know._The_reason_the_plastic_point_is_important_is~Kbr_~H~M~2~1related_to_the_sectioning_properties_of_the_wax~E_but_we_will~Kbr_~H~M~2~1come_to_that_later~3_Crystal_size_is_important_in_the_wax~Kbr_~H~M~2~1surounding_the_tissue_and_in_the_tissue_spaces~E_but_not_in_the~Kbr_~H~M~2~1tissue_per_se._Molten_wax_infiltrates_the_specimen~J_the_size~Kbr_~H~M~2~1and_shape_of_crystals_will_be_influenced_by_the_tissues_as_the~Kbr_~H~M~2~1molten_wax_solidifies_~F_i.e._crystalises._So_we_cannot_have~Kbr_~H~M~2~1~4small_crystals~4_infiltrating_although_smaller_crystals_will~Kbr_~H~M~2~1result_from_solification_in_denser_tissues.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Some_of_the_theory_behind_this_suggests_that_wax_crystalises~Kbr_~H~M~2~1first_as_flat_~4plates~E~4_the_higher_melting_point_hydrocarbons~Kbr_~H~M~2~1crystalising_first._As_successively_lower_melting_points~Kbr_~H~M~2~1deposit_further_plate_crystals~E_they_pile_up_upon_one_another.~Kbr_~H~M~2~1Distortion_due_to_these_dynamic_events_forces_the_edges_or~Kbr_~H~M~2~1corners_of_the_most_well_developed_plates_to_curl_and_roll.~Kbr_~H~M~2~1Eventually~E_that_gives_rise_to_needle_shaped_crystals~E_which~Kbr_~H~M~2~1some_~4experts~4_consider_most_ideal_for_microtomy._All_this~Kbr_~H~M~2~1will_be_contingent_upon_the_boundaries_imposed_upon_the_process~Kbr_~H~M~2~1by_cell_and_tissue_structures._During_microtomy~E_essentially~Kbr_~H~M~2~1two_types_of_forces_are_exerted_in_the_cutting_process._Flow~Kbr_~H~M~2~1shearing_and_point~Fto~Fpoint_shearing._Flow_shearing_is~E_as_you~Kbr_~H~M~2~1might_expect~E_the_smoother_and_prcedes_ahead_of_the_edge_of_the~Kbr_~H~M~2~1blade._Point_to_point_shearing_has_forces_seeking_the_line_of~Kbr_~H~M~2~1least_resistence_ahead_of_the_blade_and_these_result_in_a~Kbr_~H~M~2~1section_of_uneven_thickness_~F_not_that_you_would_notice_this~Kbr_~H~M~2~1microscopically.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Imagine_the_difference_between_cutting_through_a_jelly_and~Kbr_~H~M~2~1cutting_through_a_beefburger._Now_you_can_imagine_where_the~Kbr_~H~M~2~1importance_of_the_plastic_point_~Aas_opposed_to_the_melting~Kbr_~H~M~2~1point~B_comes_in._Additives_to_paraffin_waxes_are_intended_to~Kbr_~H~M~2~1minimise_the_point~Fto~Fpoint_shearing_and_improve_the_plastic~Kbr_~H~M~2~1flow._The_association_between_the_words_~4plastic~4_and~Kbr_~H~M~2~1~4polymers~4_should_now_be_awakening._Additives_to_paraffin_wax~Kbr_~H~M~2~1are_usually_polymers_~Aof_know_chain_length~E_for_they_are~Kbr_~H~M~2~1synthesised_exactly~B~E_with_a_major_role_in_~4harmonizing_the~Kbr_~H~M~2~1consistency~E~4_in_part_at_least_by_filling_in_beteen_the_wax~Kbr_~H~M~2~1crystals.~Kbr_~H~M~2~1~Kbr_~H~M~2~1I_use_pure_paraffin_wax_with_no_additives~E_in_the_belief_that~Kbr_~H~M~2~1proper_processing_and_a_SHARP_blade_are_the_central_features_of~Kbr_~H~M~2~1good_microtomy._~AI_just_wish_I_could_practise_as_well_as_I_can~Kbr_~H~M~2~1preach~3~B_I_have_only_ever_come_accross_one_wax_with_crystalline~Kbr_~H~M~2~1structure_significantly_different_from_others~E_and_that_is~Kbr_~H~M~2~1Ralwax~E_which_can_be_helpful_when_cutting_decalcified~Kbr_~H~M~2~1specimens~E_etc.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Xylene_substitutes~I_what_are_they~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1What_are_the_various_liquids_sold_as_substitutes_for~Kbr_~H~M~2~1xylene~E_and_are_they_really_safer_and_just_as_good~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1There_are_two_classes_of_xylene_substitutes~I_limonenes_and~Kbr_~H~M~2~1aliphatics.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Limonenes_are_prepared_by_steam_distillation_of_orange_peels.~Kbr_~H~M~2~1They_are_terpenoids_rather_similar_to_turpentine._They_are~Kbr_~H~M~2~1becoming_more_expensive_and_difficult_to_obtain._Their_great~Kbr_~H~M~2~1disadvantage_is_the_persistent_citrus_smell~E_which_many_people~Kbr_~H~M~2~1find_intolerable._They_are_difficult_to_distil._On_the_other~Kbr_~H~M~2~1hand~E_they_are_rather_minimally_toxic~E_and_are_easy_to_dispose~Kbr_~H~M~2~1of._Various_brands_are_interchangeable.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Aliphatics_are_synthetic_hydrocarbons_with_about_the_molecular~Kbr_~H~M~2~1weight_of_naphtha._They_are_odorless~E_not_very_toxic~E_and_easily~Kbr_~H~M~2~1distilled._They_are_as_difficult_to_dispose_of_as_xylene.~Kbr_~H~M~2~1~Kbr_~H~M~2~1There_are_at_least_six_brands_of_aliphatics~E_and_they_are_NOT~Kbr_~H~M~2~1interchangeable_with_each_other._They_vary_consierably_in_flash~Kbr_~H~M~2~1point~E_and_they_all_have_different_distillation_routines.~Kbr_~H~M~2~1Richard_Allen~9s_Clear~FRite_is_perhaps_the_best_known_of_them.~Kbr_~H~M~2~1Some_of_the_ones_offered_by_ma~Fand~Fpa_solvent_repackagers_are~Kbr_~H~M~2~1quite_unsatisfactory.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Bob_Richmond~E_Samurai_Pathologist~Kbr_~H~M~2~1Knoxville_TN~Kbr_~H~M~2~1~Arsrichmond~PAT~Qaol.com~B~Kbr_~H~M~2~1~Kbr_~H~M~2~1Back_to_Table_of_Contents~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Test_for_water_in_used_absolute_alcohol~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1How_can_I_determine_whether_used_~4absolute~4_alcohol_is~Kbr_~H~M~2~1still_OK_for_the_last_stage_of_dehydrating_specimens~Kbr_~H~M~2~1or_slides~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Some_people_add_anhydrous_copper_sulphate_to_the_alcohols_used~Kbr_~H~M~2~1for_processing_tissues._It_changes_colour_~Awhite_to_blue~B_in_the~Kbr_~H~M~2~1presence_of_water~E_but_this_does_not_tell_you_if_there_is_only_a~Kbr_~H~M~2~1tiny_trace_of_water_or_enough_to_make_the_alcohol_immiscible~Kbr_~H~M~2~1with_xylene.~Kbr_~H~M~2~1~Kbr_~H~M~2~1You_may_be_interested_in_a_simple_method_I_developed_for_this~Kbr_~H~M~2~1purpose._My_job_is_evaluating_histology_equipment_for_the~Kbr_~H~M~2~1Medical_Devices_Agency~E_~Aan_agency_of_the_Department_of_Health~B~E~Kbr_~H~M~2~1and_I_was_interested_in_trying_to_establish_~4carry~Fover~4_in~Kbr_~H~M~2~1processing_and_staining_instruments._I_started_off_by_adding~Kbr_~H~M~2~1known_dilutions_of_alcohol~E_drop_by_drop~E_to_different_amounts~Kbr_~H~M~2~1of_xylene~E_my_basic_thinking_being_that_water_turns_xylene~Kbr_~H~M~2~1milky~E_and_if_one_adds_enough_of_the_diluted_alcohol~E_the~Kbr_~H~M~2~1mixture_eventually_becomes_clear_again._From_this_I_developed~Kbr_~H~M~2~1the_following_method~I~Kbr_~H~M~2~1~Kbr_~H~M~2~1A_measured_5_ml_of_xylene_~Athe_5_ml_is_important~B_is_placed_in_a~Kbr_~H~M~2~150_ml_glass_beaker_and_placed_on_a_black_background._Using_a_1~Kbr_~H~M~2~1ml_plastic_pasteur~Htransfer~Hdropping_pipette~E_add_the_alcohol~Kbr_~H~M~2~1for_analysis~E_drop_by_drop_and_keep_count_of_the_number_of~Kbr_~H~M~2~1drops~E_until_you_can_just_detect_a_faint_turbidity_in_the~Kbr_~H~M~2~1xylene._Carry_on_adding_the_alcohol_to_the_xylene_until_the~Kbr_~H~M~2~1turbidity_just_clears~E_again_taking_note_of_how_many_drops_were~Kbr_~H~M~2~1needed.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Using_known_dilutions_of_alcohol~E_I_was_able_to_set_up_and~Kbr_~H~M~2~1standardise_the_method_and_obtain_reproduceable_results~Kbr_~H~M~2~1consistently._The_method_was_not_sensitive_enough_to_detect_the~Kbr_~H~M~2~1water_in_99~7_or_98~7_alcohol.~Kbr_~H~M~2~1~Kbr_~H~M~2~197~7_~L_5_drops_to_turn_xylene_milky~E_10_drops_to_clear_the_mixture~Kbr_~H~M~2~196~7_~L_4_drops_to_turn_xylene_milky~E_14_drops_to_clear_the_mixture~Kbr_~H~M~2~195~7_~L_3_drops_to_turn_xylene_milky~E_34_drops_to_clear_the_mixture~Kbr_~H~M~2~194~7_~L_3_drops_to_turn_xylene_milky~E_74_drops_to_clear_the_mixture~Kbr_~H~M~2~193~7_~L_3_drops_to_turn_xylene_milky~E_83_drops_to_clear_the_mixture~Kbr_~H~M~2~192~7_~L_3_drops_to_turn_xylene_milky~E_98_drops_to_clear_the_mixture~Kbr_~H~M~2~191~7_~L_3_drops_to_turn_xylene_milky~E_140_drops_to_clear_the_mixture~Kbr_~H~M~2~190~7_~L_3_drops_to_turn_xylene_milky~E_204_drops_to_clear_the_mixture~Kbr_~H~M~2~1~Kbr_~H~M~2~1You_would_have_to_initially_set_up_your_own_range_of_standard~Kbr_~H~M~2~1dilutions_with_the_particular_alcohol_used_in_your_laboratory~Kbr_~H~M~2~1for_the_sake_of_accuracy._The_1_ml_plastic~Kbr_~H~M~2~1pasteur~Htransfer~Hdropping_pipettes~E_they_can_even_be_called~Kbr_~H~M~2~1pastettes~E_should_be_held_vertically_to_standardise_the_size_of~Kbr_~H~M~2~1the_drops~E_and_I_tried_to_use_the_same_brand_each_time.~Kbr_~H~M~2~1~Kbr_~H~M~2~1This_is_a_simple_method~E_and_quick_to_do~E_although_I_should~Kbr_~H~M~2~1think_the_method_would_give_the_Biochemists_the_shudders._It~Kbr_~H~M~2~1could_help_to_prolong_the_life_expectancy_of_the_alcohols_used~Kbr_~H~M~2~1in_processors.~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~1~Kbr_~H~M~2~undefined_Molecular_sieves_for_making_anhydrous_solvent~Kbr_~H~M~2~1~Kbr_~H~M~2~1Question.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Which_type_of_molecular_sieves_are_used_for_making_anyydrous~Kbr_~H~M~2~1acetone_or_alcohol~E_and_how_much_should_I_put_in_the_bottle~N~Kbr_~H~M~2~1~Kbr_~H~M~2~1Answer.~Kbr_~H~M~2~1~Kbr_~H~M~2~1The_molecular_sieve_to_use_for_acetone_is_type_3A~E_mesh_8~F12.~Kbr_~H~M~2~1EM_Science_Catalog_~5_MX1583L~H1_for_500_g_or_~H3_for_2kg.~Kbr_~H~M~2~1~Kbr_~H~M~2~1Before_using_a_molecular_sieve~E_you_first_have_to_determine~Kbr_~H~M~2~1which_one_to_use._Type_3A_if_for_unsaturated_h