β-Defensin 22 is a major component of the mouse sperm glycocalyx

Surface components of sperm isolated from the cauda epididymides were stabilized by whole sperm fixation for immunization of rabbits. The resulting immunoglobulins (Igs) recognized a single protein of 130 kDa (non-reduced) or 54–57 kDa (reduced) on western blots of cauda sperm. Igs recognized the same 54–57 kDa protein band on whole tissue blots of the corpus and cauda epididymidis and vas deferens. No immunoreactive bands were detected on blots of the prostate, seminal vesicles, testes, caput epididymis, or any of various non-reproductive tissues. Removal of sperm from the vas deferens prior to blotting eliminated the detection of the sperm antigen. Antibodies raised to synthetic peptides, identical in amino acid sequence to two unique spans of DEFB22, recognized the same 130/54–57 kDa antigen on western blots of both caudal sperm and the purified antigen isolated with the anti-sperm Ig. From indirect immunofluorescence, both the anti-sperm and anti-peptide Igs appeared to localize to the entire sperm surface, a pattern confirmed at the ultrastructural level. Real-time PCR identified the corpus epididymides as the major site of expression of DEFB22, with negligible expression in the testes, caput epididymides, and vas deferens. Immunostaining of epididymal sections showed DEFB22 being released into the lumen at the distal caput/proximal corpus, with sperm becoming intensely coated with DEFB22 as they reached the distal corpus. Most uterine sperm recovered from mice 4 h following copulation exhibited DEFB22 coating the entire sperm surface. By contrast, some sperm recovered from the oviduct and cumulus extracellular matrix showed loss of DEFB22 from the sperm head.

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Characterization of a monoclonal antibody against active pectate lyase from Erwinia carotovora

Canadian Journal of Microbiology ◽

10.1139/m89-105 ◽

1989 ◽

Vol 35 (6) ◽

pp. 651-655 ◽

Cited By ~ 5

Author(s):

L. J. Ward ◽

S. H. De Boer

Keyword(s):

Monoclonal Antibody ◽

Active Form ◽

Sodium Dodecyl ◽

Pectate Lyase ◽

Erwinia Carotovora ◽

Protein Band ◽

Major Protein ◽

Western Blots ◽

Diverse Range ◽

Purified Antigen

A monoclonal antibody (2E2) produced against pectate lyase from Erwinia carotovora ssp. carotovora reacted with a 41-and a 44-kilodalton protein on Western blots of concentrated Erwinia culture supernatants resolved by sodium dodecyl sulphate – polyaerylamide gel electrophoresis. It was unequivocally shown that monoclonal 2E2 reacted with an active form of pectate lyase by affinity purifying the antigen with the monoclonal. The affinity-purified antigen was enzymatically active and moved as a single protein band in a nonequilibrium isoelectric focusing gel. Monoclonal 2E2 reacted with the pectate lyases of a diverse range of E. carotovora ssp. carotovora, ssp. atroseptica, and ssp. betavasculorum strains, as well as with one of three strains of E. chrysanthemi. The electrophoretic mobility of the major protein (44 kilodaltons) that reacted with 2E2 was identical within a subspecies but differed among subspecies.Key words: Erwinia carotovora, pectate lyase, monoclonal antibody.

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Glycoconjugates on the surface of epididymal spermatozoa in a marsupial, the brushtail possum, Trichosurus vulpecula

Reproduction ◽

10.1530/rep.0.1220165 ◽

2001 ◽

pp. 165-176 ◽

Cited By ~ 6

Author(s):

NJ Cooper ◽

RV McClean ◽

CM Leigh ◽

WG Breed

Keyword(s):

Fluorescein Isothiocyanate ◽

Surface Proteins ◽

Brushtail Possum ◽

Trichosurus Vulpecula ◽

Western Blots ◽

Sperm Surface ◽

Lectin Staining ◽

Epididymal Spermatozoa ◽

Cauda Epididymidis ◽

Caput Epididymidis

Variation in localization and distribution of saccharides on the sperm surface of a marsupial, the brushtail possum, Trichosurus vulpecula, was compared between spermatozoa from the caput and cauda epididymides. Spermatozoa were subjected to the following treatments: (i) unfixed and fixed spermatozoa were stained with fluorescein-labelled lectins; (ii) unfixed spermatozoa were incubated with lectins for determination of agglutination; and (iii) spermatozoa were incubated with detergent to remove the plasmalemma, the glycoproteins were separated on SDS-PAGE and western blots were stained with biotinylated lectins. Many of the fluorescein isothiocyanate (FITC)-labelled lectins bound selectively to the sperm surface, and marked differences were found in lectin staining affinity between caput and cauda epididymal spermatozoa. Incubation of spermatozoa from the cauda epididymidis with neuraminidase reversed many of the differences in staining of the cauda epididymal spermatozoa, indicating masking of some terminal saccharides by sialic acid. Agglutination of spermatozoa from the caput epididymidis occurred after incubation with Concanavalin A (ConA) and soybean agglutinin (SBA), but agglutination was less extensive for spermatozoa from the cauda epididymidis. Western blot analysis indicated several ConA-positive bands in caput sperm extracts, but fewer positive bands in the cauda sperm extracts, whereas SBA stained four bands from caput but none from the cauda epididymal spermatozoa. These results demonstrate extensive glycosylation of the surface proteins of spermatozoa from the caput epididymidis and significant differences in spermatozoa from the cauda epididymidis. In general, the findings indicate similar glycosylation of the surface of marsupial spermatozoa to those from eutherian mammals despite marked differences in their morphology and early divergence of marsupials from eutherian mammals. It would appear that this situation differs markedly from that in sub-mammalian vertebrates.

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368. A direct control of the prostate and seminal vesicles via the vas deferens

Journal of Steroid Biochemistry ◽

10.1016/0022-4731(74)90513-5 ◽

1974 ◽

Vol 5 (4) ◽

pp. 383 ◽

Cited By ~ 2

Author(s):

C.G. Pierrepoint ◽

P. Davies ◽

Barbara A. John

Keyword(s):

Vas Deferens ◽

Seminal Vesicles ◽

Direct Control

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Vas deferens invasion: A neglected issue in the sampling of radical prostatectomy materials

Canadian Urological Association Journal ◽

10.5489/cuaj.1802 ◽

2014 ◽

Vol 8 (7-8) ◽

pp. 554 ◽

Cited By ~ 3

Author(s):

Nuri Yigit ◽

Yildirim Karslioglu ◽

Bulent Kurt

Keyword(s):

Lymph Node ◽

Radical Prostatectomy ◽

Lymph Node Metastasis ◽

Vas Deferens ◽

Prognostic Significance ◽

Seminal Vesicles ◽

Node Metastasis ◽

Increased Risk ◽

Staging Systems ◽

Independent Prognostic Significance

A radical prostatectomy affects the prostate, bilateral seminal vesicles (SV), and the distal parts of the bilateral vasa deferentia (VD). SV invasion (SVI) is associated with an increased risk of lymph node metastasis and recurrence. However, the significance of VD invasion (VDI), either with or without the involvement of their surgical margins, has not been fully appreciated. We think VDI might have an independent prognostic significance, as does SVI, and should be incorporated into the pathology guidelines and the staging systems of prostatic adenocarcinoma. Our case illustrates this.

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Purification of p-hydroxyphenylpyruvate hydroxylase from rat liver—requirements for cofactors

Canadian Journal of Biochemistry ◽

10.1139/o76-061 ◽

1976 ◽

Vol 54 (5) ◽

pp. 423-431 ◽

Cited By ~ 5

Author(s):

Kun-Tsan Lin ◽

John C. Crawhall

Keyword(s):

Molecular Weight ◽

Enzyme Activity ◽

Rat Liver ◽

Gel Filtration ◽

Protein Band ◽

Final Concentration ◽

Assay Method ◽

Enzyme Extract ◽

Crude Enzyme Extract ◽

Single Protein

Theenzyme p-hydroxyphenylpyruvate hydroxylase (EC 1.13.11.27)from rat liver was studied with the assay method which measures the release of 14CO2 from p-hydroxyphenyl [carboxyl-,14C]pyruvate. Extensive dialysis of the crude enzyme extract against Tris buffer or purification involving ammonium sulfate, gel filtration, and ion exchange results in loss of enzyme activity that can be reactivated by Fe2+, dichlorophenolindophenol, and various other agents. The effect of these activators depends critically on their final concentration in the assay media.A 70-fold purification of the enzyme fraction yielded a preparation which behaved as a single protein band in Sephadex G-150. It had an isoelectric point at 5.85 and molecular weight of 63 000. The enzyme obtained appears to be different in some respects from those described by other workers from the liver of dog, human, chicken, and frog.

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SCANNING ELECTRON MICROSCOPY OF HUMAN VAS DEFERENS AND SEMINAL VESICLES

Three Dimensional Microanatomy of Cells and Tissue Surfaces ◽

10.1016/b978-0-444-00607-3.50024-x ◽

1981 ◽

pp. 299-310 ◽

Cited By ~ 4

Author(s):

GIOVANNI E. ORLANDINI ◽

PAOLO PACINI ◽

MASSIMO GULISANO

Keyword(s):

Electron Microscopy ◽

Scanning Electron Microscopy ◽

Vas Deferens ◽

Seminal Vesicles ◽

Scanning Electron

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[3H]thymidine uptake by the epididymis, seminal vesicles and prostate gland during postnatal development of the rat

Reproduction Fertility and Development ◽

10.1071/rd9910313 ◽

1991 ◽

Vol 3 (3) ◽

pp. 313 ◽

Cited By ~ 3

Author(s):

S Sujarit ◽

RC Jones

Keyword(s):

Postnatal Development ◽

Similar Pattern ◽

Initial Segment ◽

Prostate Gland ◽

Seminal Vesicles ◽

Ventral Prostate ◽

Thymidine Uptake ◽

Low Levels ◽

Cauda Epididymidis

The uptake of [3H]thymidine by the epididymis, ventral prostate gland and seminal vesicles was determined in vivo for rats aged 15, 20, 25, 30, 35, 45 and 55 days. The pattern of uptake varied considerably between organs and generally was different from patterns of growth measured as mass or ratio of mass of DNA:tissue. The 'initial segment' of the epididymis and caput and corpus epididymidis showed a similar pattern of [3H]thymidine uptake, being greatest in 15-day-old animals and declining thereafter. On Day 15 the cauda epididymidis had a lower uptake than more proximal regions of the epididymis, but it subsequently showed two significant peaks of increased uptake on Days 25-30 and Day 45. The uptake by the seminal vesicles was high on Day 15, fell to low levels on Day 20, increased considerably from Days 20 to 35, then gradually decreased from Day 35 to 55. The uptake by the prostate gland was a little lower than by the seminal vesicles on Days 15 and 20, then reduced to about the same level as non-reproductive tissues.

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Electrogenic 2 Na/1 H antiport in crustacean epithelium is inhibited by a monoclonal antibody

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.1993.264.4.r804 ◽

1993 ◽

Vol 264 (4) ◽

pp. R804-R810

Author(s):

H. Gert de Couet ◽

L. Busquets-Turner ◽

A. Gresham ◽

G. A. Ahearn

Keyword(s):

Brush Border ◽

Membrane Vesicles ◽

Structural Similarity ◽

Protein Band ◽

Monovalent Cation ◽

Cation Binding ◽

Western Blots ◽

System A ◽

Published Evidence

We have previously published evidence that suggests that Na/H exchange in crustacean and echinoderm epithelia occurs by an electrogenic antiporter protein with two external cation binding sites that accommodate Na, amiloride, or Ca and display a 2:1 monovalent cation antiport stoichiometry. The present study is an initial investigation into the molecular biology of this invertebrate electrogenic exchanger to ascertain its structural similarity to the analogous vertebrate electroneutral antiport system. A panel of monoclonal antibodies was prepared against components of lobster hepatopancreatic epithelial brush-border membranes and assayed immunohistochemically and by Western blotting. The antibodies were tested further in functional assays for their ability to interfere with electrogenic 2 Na/1 H antiport in isolated hepatopancreatic brush-border membrane vesicles. One cell line was identified producing an antibody that significantly inhibited the electrogenic exchange of cations by these membrane preparations and recognized a single protein band on Western blots of hepatopancreas, antennal gland, and gill epithelia corresponding to a molecular mass of 185 kDa. The existence of such an antibody probe may facilitate the purification of the electrogenic antiporter under denaturing conditions, in in vitro expression systems, or in prokaryotic expression libraries.

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Nanomaterials Enhanced Gene Expression in Yeast Cells

Journal of Nanomaterials ◽

10.1155/2008/391497 ◽

2008 ◽

Vol 2008 ◽

pp. 1-9 ◽

Cited By ~ 1

Author(s):

Su-Fang Chien ◽

Shi-Hui Chen ◽

Chhiu-Tsu Lin

Keyword(s):

Gene Expression ◽

Ag Nanoparticles ◽

Product Yield ◽

Protein Band ◽

Yeast Cells ◽

Vis Spectroscopy ◽

Yeast Chromosome ◽

Single Protein ◽

Enhance Gene Expression ◽

Metal Nanomaterials

Metal nanomaterials are shown to enhance gene expression for rice -galactosidase gene (-Gal) in yeast cells. Au and Ag nanoparticles and their nanocomposites, silica-Au and silica-Ag, were prepared and characterized by UV-vis spectroscopy and TEM technique. The rice -galactosidase gene was cloned into the yeast chromosome, where the cloned cells were precultured and induced into a medium containing each of the testing nanomaterials. The nanomaterials were observed to incorporate inside the cells, and no cell death has been detected during the course of gene expression. The enzyme activity was determined by a synthetic substrate, p-nitrophenyl--D-galctopyranoside, and the yellow product yield was recorded in a spectrophotometer at 400 nm. When Au and Ag nanoparticles were incorporated with the culture, a 3–5 fold enhancement in -galactosidase was observed for intracellular activity as well as the secreted activity into the medium. The secreted protein was analyzed to have a pure form and displayed as a single protein band in the SDS-gel electrophoresis. The effects of size and chemical nature of nanomaterials on gene expression for the rice -galactosidase gene in yeast cells are discussed.

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