COMBINED USE OF FLUORESCENT PEANUT AGGLUTININ LECTIN AND HOECHST-33258 TO MONITOR THE ACROSOMAL STATUS AND VITALITY OF HUMAN SPERMATOZOA

被引：53

作者：

MORTIMER, D

CURTIS, EF

CAMENZIND, AR

机构：

[1] Gamete Biology Research Unit, Reproductive Medicine Research Group, University of Calgary Health Sciences Centre, Calgary, AB T2N 4N1

来源：

HUMAN REPRODUCTION | 1990年 / 5卷 / 01期

基金：

英国医学研究理事会;

关键词：

Acrosome; Human sperm; Lectin; Spermatozoa; Vitality;

D O I：

10.1093/oxfordjournals.humrep.a137050

中图分类号：

R71 [妇产科学];

学科分类号：

100211 ;

摘要：

Fluorescein-conjugated peanut agglutinin (PNA) lectin-labelling is an established procedure for assessing the status of the human sperm acrosome. However, unlike the triple-stain technique, PNA-labelling does not provide a simultaneous assessment of cellular vitality. We have therefore evaluated the use of the fluorescent dye Hoechst 33258 (H33258) as a vital stain for use in combination with PNA-labelling. Human sperm populations were stained for 1 min with 1 μg/ml H33258 in culture medium and then washed through 2.0 (w/v) polyvinylpyrollidone columns and air-dried onto microscope slides. H33258 was found to provide vitality assessments comparable to those obtained using the standard eosin-exclusion method. However, best results were obtained with an ethanol fixation step between air-drying and PNA-labelling. This vitality assessment was found to be more reliable than that provided by Trypan blue staining under conditions equivalent to the triple-stain technique. There was no alteration of PNA-labelling due to the H33258 although ethanol fixation actually provided more uniform PNA-labelling than previously obtained without ethanol fixation. Consequently, we have stopped using the triple-stain technique for assessing human sperm acrosome reactions and now use the H33258/PNA procedure routinely. © 1990 Oxford University Press.

引用

页码：99 / 103

页数：5

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