白蛋白XY精蟲分離原理

白蛋白精蟲分離原理為Y精蟲在白蛋白中游的比X精蟲快
但FISH顯示XY精蟲比例並不明顯改變
可能機轉為白蛋白精蟲分離較容易造成X精蟲去活化
使的Y精蟲受孕機率提高

http://humrep.oxfordjournals.org/content/13/1/146.long

Hum Reprod. 1998 Jan;13(1):146-9.

Experiences in Hong Kong with the theory and practice of the albumin column method of sperm separation for sex selection.

Rose GA, Wong A.

Source

Gender Choice Centre, Central, Hong Kong.

Erratum in

• Hum Reprod 1998 May;13(5):1414.

Abstract

Controversy still surrounds the human serum albumin (HSA) method for separation of X- and Y-bearing human spermatozoa. There is doubt about whether the procedure does enrich sperm samples for the chosen sex chromosome. We have applied the HSA separation method in a clinic in Hong Kong, using the method as described by Ericsson et al. [Nature, 246, 421-424 (1973)] taking care to keep the sperm recovery to <5% of the initial number. Aliquots of separated spermatozoa were examined for X- and Y-bearing spermatozoa by fluorescent in-situ hybridization (FISH) using appropriate DNA probes. Of 18 couples wanting boys, 13 had single boys, one had twin boys, and one had twins comprising one boy and one girl. Only three single girls were born. This success rate of 83% is significantly different (P < 0.001) from the usual expected ratio. There were four miscarriages, one in the third and one in the fourth week of pregnancy. The times of the others are not definitely known, but are thought to have occurred early in pregnancy. We lack information on three couples. The FISH procedure showed no change in the normal and equal numbers of X- and Y-bearing spermatozoa after the HSA separation procedure. This study confirmed that the HSA sperm separation method can bias the number of babies in favour of males. However, the theory that it does so by enriching the sperm samples with Y-bearing spermatozoa appears to be incorrect and some other theory has to be postulated. It is tentatively proposed that passage through the HSA inactivates X-bearingspermatozoa more than Y-bearing spermatozoa, even though this is not apparent simply on inspection of sperm motility.

PIP:

The ability of the human serum albumin (HSA) method for separation of X- and Y-bearing human spermatozoa to enrichsperm samples for the chosen sex chromosome was investigated in 112 couples inseminated at the Gender Choice Center in Hong Kong. Sperm recovery was limited, as specified by Ericsson, to less than 5% of the initial number. Aliquots of separated spermatozoa were examined for X- and Y-bearing spermatozoa by fluorescent in situ hybridization with use of DNA probes. All 30 women who conceived during the study period wanted boys. Of 18 of these women for whom results were available, 13 had single boys, 1 had twin boys, 1 had male-female twins, and 3 had single girls. The 83% success rate recorded in this series is significantly different (p 0.001) from the expected ratio. There were 4 miscarriages, all of which occurred in the first trimester, and 2 ectopic pregnancies. These findings indicate that the HSA sperm separation method can indeed create a bias toward the birth of a male infant. However, since there were no changes in the normal and equal numbers of X- and Y-bearing spermatozoa after the HSA procedure, the theory that the procedure enriches sperm samples with Y-bearing spermatozoa appears to be incorrect. It is tentatively postulated that passage through the HSA inactivates X-bearing spermatozoa more than Y-bearing spermatozoa.

The supernatant

was poured off and the pellet diluted with Tyrode solution to give a

sperm concentration of ~203106/ml. 10% HSA solution was prepared

from 20% HSA in the separation kit. The number of columns prepared

depended upon the volume of resuspended sperm samples. 1 ml of

10% HSA solution was gently layered into each column. 0.5 ml of

sperm sample was gently layered on top of each 10% HSA layer.

After 45 min 0.5 ml of the top sperm layer was aspirated and the

10% HSA from each column were pooled. Total sperm count,

percentage motility and amount of debris were checked in the pooled

top sperm layers and the pooled 10% HSA layers were centrifuged

at 600 g for 10 min. The supernatant was removed and the pellet

diluted with Tyrode’s solution. Columns with 1.0 ml of 12% HSA

on the top of 0.5 ml of 20% HSA were prepared. 0.5 ml of resuspended

spermatozoa gently layered on top of each 12% HSA layer. After 45

min 0.5 ml of the top layers were aspirated and pooled. The 12 and

20% HSA layers were also aspirated and pooled separately. Total

sperm counts, percentage motilities and amount of debris were

checked in each layer. The pooled 20% HSA layers were diluted

with equal volume and centrifuged at 600 g for 10 min. The sperm

pellet was removed and the pellet resuspended in 0.5 ml of Tyrode’s

solution for artificial insemination. This procedure was found to give

yields of motile spermatozoa of ,10% in accordance with the

recommendation of Pyrzak (1994) and Ericsson (1994).