1. Fertility and Sterility
Identification of human sperm proteins involved in the interaction with homologous zona pellucida 
Andrea Lasserre, Ph.D., Fernanda González-Echeverría, M.Sc., Carolina Moules, M.Sc., Jorge G Tezón, Ph.D., Patricia V Miranda, Ph.D., Mónica H Vazquez-Levin, Ph.D. 
Fertility and Sterility 
Volume 79, Pages (June 2003)
DOI: /S (03)

2. FIGURE 1
HSE proteins and immunoreactivity of anti-HSE. (A) High salt extract (HSE) components were separated in 8% SDS-PAGE and stained with CBB. Samples were reduced by adding 5% 2-β-mercaptoethanol before electrophoresis. MWM: molecular weight markers (kd). This experiment was performed at least five times obtaining similar results. A typical experiment is shown. (B) Protein extracts were subjected to 8% SDS-PAGE, followed by Western immunoblotting with anti-HSE. This experiment was Performed at least five times obtaining similar results. A typical experiment is shown. (Lane a) High salt extract (HSE) proteins. (Lane b) Proteins from human epididymis. (Lane c) Cauda epididymal plasma (CEP) proteins. (Lane d) Proteins from human testis. (Lane e) Seminal plasma proteins.
Lasserre. Human sperm proteins and zona pellucida interaction. Fertil Steril 2003.
Fertility and Sterility  , DOI: ( /S (03) )

3. FIGURE 2
Immunolocalization of p49 and p66 in spermatozoa. Immunocytochemical analysis of ejaculated (a, c, e) and 4-hour capacitated (b, d, f) spermatozoa stained with anti-HSE (a, b), anti-p66 (c, d), and anti-p49 (e, f) antibodies. Staining performed with preimmune serum (g: bright-field, h: immunofluorescence) (magnification, ×400).
Lasserre. Human sperm proteins and zona pellucida interaction. Fertil Steril 2003.
Fertility and Sterility  , DOI: ( /S (03) )

4. FIGURE 3
Immunoreactivity of anti-p49 and anti-p66 antibodies for proteins from the male reproductive tract. Protein extracts from the male reproductive tract were subjected to 8% SDS-PAGE followed by Western immunoblotting with anti-p66 (A) and anti-p49 (B) antibodies. (Lane a) High salt extract (HSE) proteins. (Lane b) Proteins from human epididymis. (Lane c) Cauda epididymal plasma (CEP) proteins. (Lane d) Proteins from human testis. (Lane e) Seminal plasma proteins.
Lasserre. Human sperm proteins and zona pellucida interaction. Fertil Steril 2003.
Fertility and Sterility  , DOI: ( /S (03) )

5. FIGURE 4
Immunolocalization of p66 and p49 in ejaculated and 4-hour capacitated spermatozoa. Assessment of sperm staining patterns with anti-p66 and anti-p49 antibodies was done in at least 200 spermatozoa from four semen samples.
Lasserre. Human sperm proteins and zona pellucida interaction. Fertil Steril 2003.
Fertility and Sterility  , DOI: ( /S (03) )

6. FIGURE 5
Effect of anti-p66 and anti-p49 antibodies on sperm–ZP interaction. Spermatozoa were preincubated with 100 μg/mL of a mixture of anti-p49 + anti-p66 antibodies or preimmune serum (control) and washed, followed by incubation with homologous ZP using the HZA. In all cases, spermatozoa tightly bound to the outer face of each HZ were counted. Results show the number of spermatozoa bound to each HZ (n = 6) (mean ± SD).
Lasserre. Human sperm proteins and zona pellucida interaction. Fertil Steril 2003.
Fertility and Sterility  , DOI: ( /S (03) )

7. FIGURE 6
Effect of sperm proteins p66 and p49 and HSA on sperm–ZP interaction. Spermatozoa were incubated with homologous ZP in the HZA in the presence of 10 μg/mL of sperm proteins p66 (A) or p49 (B). In all cases, spermatozoa tightly bound to the outer face of each HZ were counted. Results show the number of spermatozoa bound to each HZ (n = 5) (mean ± SD).
Lasserre. Human sperm proteins and zona pellucida interaction. Fertil Steril 2003.
Fertility and Sterility  , DOI: ( /S (03) )

8. FIGURE 7
Immunolocalization of p66 in human epididymis and testis. Tissue sections from distal caput (a, b, c), corpus (d, e, f) and cauda (g, h, i) epididymis, and testis (j, k, l) were immunostained with anti-p66 (a, d, g, j) and preimmune serum (b, e, h, k). Sections stained with hematoxylin (c, f, i, l) are also included (magnification, ×100).
Lasserre. Human sperm proteins and zona pellucida interaction. Fertil Steril 2003.
Fertility and Sterility  , DOI: ( /S (03) )