PMID- 15867004
OWN - NLM
STAT- MEDLINE
DCOM- 20050921
LR  - 20191210
IS  - 0196-3635 (Print)
IS  - 0196-3635 (Linking)
VI  - 26
IP  - 3
DP  - 2005 May-Jun
TI  - Variability and laboratory factors affecting the sperm chromatin structure assay 
      in human semen.
PG  - 360-8
AB  - During the past decade, the sperm chromatin structure assay (SCSA) has become an 
      important tool for assessing semen quality in the human andrology laboratory. The 
      SCSA uses the metachromatic properties of the fluorescent dye acridine orange 
      (AO) in combination with flow cytometry to determine the sperm DNA susceptibility 
      to denaturation in situ. The objective of this study was to evaluate laboratory 
      factors affecting the SCSA and the variation between replicates. Semen ejaculates 
      from 3 healthy volunteers were analyzed using the SCSA protocol as described by 
      Evenson and Jost (2000), determining the X-mean, Y-mean, DNA fragmentation index 
      (DFI), standard deviation of DFI (SD-DFI), and high DNA stainability (HDS). In 
      experiment 1, the effects of thawing time, time of day, day, laboratory 
      technician, donor, and incubation period before analysis were investigated. In 
      experiment 2, the effects of sheath fluid, AO equilibration buffer, day, 
      laboratory technician, donor, and incubation period before analysis were 
      investigated. A significant difference was found between the 3 donors with 
      respect to the X-mean, Y-mean, DFI, SD-DFI, and HDS. It was shown that incubation 
      of the semen samples on ice postthaw had a significant effect on the X-mean, 
      Y-mean, DFI, and SD-DFI. The laboratory technician conducting the analysis 
      accounted for up to 15.4% for the variation of the SCSA measurements. The time of 
      day affected the variation for the Y-mean (23.5% of the total variation of the 
      Y-mean), and the day affected the variation for the X-mean (82.8% of the total 
      variation of the X-mean). Incubation on ice for 5 to 25 minutes postthaw had a 
      significant effect on the DFI and SD-DFI in both experiments. This study shows 
      that several protocol steps in the SCSA affect the results obtained from the 
      assay. Precise protocol description and standardization of the SCSA are therefore 
      essential to achieve high agreement within and between different laboratories.
FAU - Boe-Hansen, Gry B
AU  - Boe-Hansen GB
AD  - The Royal Veterinary and Agricultural University, Department of Large Animal 
      Sciences, Section for Veterinary Reproduction and Obstetrics, Dyrlaegevej 68, 
      1870 Frederiksberg C, Denmark. gbh@kvl.dk
FAU - Ersboll, Annette K
AU  - Ersboll AK
FAU - Christensen, Preben
AU  - Christensen P
LA  - eng
PT  - Evaluation Study
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Validation Study
PL  - United States
TA  - J Androl
JT  - Journal of andrology
JID - 8106453
RN  - 0 (Chromatin)
SB  - IM
MH  - Chromatin/*ultrastructure
MH  - Clinical Laboratory Techniques/standards
MH  - DNA Fragmentation
MH  - Flow Cytometry
MH  - Freezing
MH  - Humans
MH  - Male
MH  - Reproducibility of Results
MH  - Semen/cytology
MH  - Spermatozoa/*ultrastructure
MH  - Time Factors
EDAT- 2005/05/04 09:00
MHDA- 2005/09/22 09:00
CRDT- 2005/05/04 09:00
PHST- 2005/05/04 09:00 [pubmed]
PHST- 2005/09/22 09:00 [medline]
PHST- 2005/05/04 09:00 [entrez]
AID - 26/3/360 [pii]
AID - 10.2164/jandrol.04056 [doi]
PST - ppublish
SO  - J Androl. 2005 May-Jun;26(3):360-8. doi: 10.2164/jandrol.04056.