Revision 21_Sep_2015
35
User Guide Version 2.60 I-button WHO 5th
APPENDIX 4: Counting Cells using the SQA-V Visualization System
1.
Follow the WHO Manual instructions for semen sample collection and preparation. Thoroughly mix the
sample before step #2.
2.
Pipette 10uL of the semen sample onto a standard slide and cover with a 22x22 mm cover slip. Prepare a
new slide if air bubbles or liquid spillage occurs.
3.
Load the slide into the slide adaptor and then insert the slide adaptor into the SQA-V visualization
chamber. (Refer to the SQA-V User Guide APPENDIX 3: Using Standard Slides in the Visualization System
for details).
4.
Press the ZOOM-OUT button on the SQA-V keypad all the way to set the magnification to x300.
5.
Set the: BRIGHTNESS, CONTRAST & COLOR knobs of the video display:
a.
COLOR knob: Turn clockwise to the end (maximum color),
b.
CONTRAST: Turn counterclockwise to the end (maximum contrast),
c.
BRIGHTNESS knob: Turn clockwise from the darkest setting until the background is light (not
maximum!).
6.
Adjust the focus knob to maximize the image: Turn clockwise all the way. Then turn counterclockwise
until a clear image appears on the screen.
7.
Go to V-Sperm and click on the
Real Time Video
button. FREEZE the image.
8.
The screen of both the SQA-V and the V-Sperm is divided into a grid containing 20-distinct squares (see
below).
9.
Each spermatozoon seen on the ENTIRE 20-square grid is 1 Million/ml of sperm concentration. FOR
EXAMPLE: In the grid above, there are 7 spermatozoa in each cell of the grid. 7 (spermatozoa) X 20
(cells) = 140 M/ml sperm concentration for this sample.
10.
To count a minimum of 200 cells (per WHO), turn the silver knob of the slide adaptor and a new field of
view will be displayed in the grid.
11.
When viewing multiple fields, divide the final count by the number of screens (fields of view) counted. For
example, if two of the screens above are counted there would be a total number of 280 sperm cells so
the sperm concentration will be: 280 ÷ 2 = 140 M/ml.
12.
Refer to table 2.2 of the WHO Manual 5
th
Edition to determine if the duplicate counts are acceptable.
