A comparison of methods for preparing enriched populations of bovine spermatogonia

Herrid, Muren, Davey, Rhonda J., Hutton, Keryn, Colditz, Ian G. and Hill, Jonathan R. (2009) A comparison of methods for preparing enriched populations of bovine spermatogonia. Reproduction, Fertility and Development, 21 3: 393-399. doi:10.1071/RD08129


Author Herrid, Muren
Davey, Rhonda J.
Hutton, Keryn
Colditz, Ian G.
Hill, Jonathan R.
Title A comparison of methods for preparing enriched populations of bovine spermatogonia
Journal name Reproduction, Fertility and Development   Check publisher's open access policy
ISSN 1031-3613
Publication date 2009-11
Sub-type Article (original research)
DOI 10.1071/RD08129
Volume 21
Issue 3
Start page 393
End page 399
Total pages 7
Editor Tony Flint
Place of publication East Melbourne, Victoria, Australia
Publisher CSIRO in co-operation with the Australian Academy of Science
Collection year 2010
Language eng
Subject C1
0707 Veterinary Sciences
Formatted abstract
The objective of the present study was to identify an efficient and practical enrichment method for bovine type A spermatogonia. Four different enrichment methods were compared: differential plating on laminin- or Datura stramonium agglutinin (DSA)-coated flasks, percoll-gradient isolation, magnetic-activated cell sorting (MACS) and fluorescence-activated cell sorting (FACS). The isolated cells were characterised with Dolichos biflorus agglutinin (DBA) lectin staining for type A spermatogonia and vimentin-antibody staining for Sertoli cells. A 2 × 2 factorial design was used to investigate the enrichment efficiency on laminin and DSA. In the laminin-enrichment groups, 2 h incubation in plates coated with 20 μg mL–1 laminin yielded a 3.3-fold increase in DBA-positive cells in the adherent fraction, while overnight incubation in flasks coated with 20 μg mL–1 DSA produced a 3.6-fold increase in the non-adherent fraction. However, the greatest enrichment (5.3-fold) of DBA-positive cells was obtained after 2 h incubation in control flasks (coated with bovine serum albumin). Percoll-gradient centrifugation yielded a 3-fold increase in DBA-positive cells. MACS results showed a 3.5- to 5-fold enrichment while FACS produced a 4-fold increase in DBA-positive cells. It is concluded that differential plating is a better method of recovering large numbers of type A spermatogonia for germ cell transplantation, while MACS or FACS can provide highly enriched viable type A spermatogonia for in vitro culture. Further, the combination of differential plating and other enrichment techniques may increase the purification efficiency of type A spermatogonia.
Keyword enrichment
isolation
stem cells
Q-Index Code C1
Q-Index Status Confirmed Code

Document type: Journal Article
Sub-type: Article (original research)
Collections: 2010 Higher Education Research Data Collection
School of Veterinary Science Publications
 
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Citation counts: TR Web of Science Citation Count  Cited 31 times in Thomson Reuters Web of Science Article | Citations
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Created: Fri, 18 Jun 2010, 10:49:46 EST by Matthew Lamb on behalf of Faculty Of Nat Resources, Agric & Veterinary Sc