
Leja Slide Semen Analysis ChamberStandard Count Disposable Analysis Chambers Cleaner, higherquality glass and a unique design deliver more
uniform semen samples and more accurate counts! Clumping of cells makes accurate counting tough. This clumping may be caused when cells gather, and particulate around air bubbles,
pitting in the glass or particulate contamination. Only Standard
Count Analysis Chambers eliminates all three causes of clumping,
Instructions for use loading the Standard Count ChamberUsing a positive displacement pipet, place approximately 3 to 5 ul of sample into one of the two clear zones labeled SAMPLE A and SAMPLE B. Allow a few seconds for the sample to completely load into the analysis area, then remove any excess fluid remaining in the loading zone. Perform the analysis near the center of a large open area of the Standard Count Chamber. Automated AnalysisThe Standard Count is compatible with all CASA systems. Because there is no counting grid etched into the chamber, you may select fields for analysis from any part of the chamber.
Manual AnalysisNOTE: If you are currently using a disposable counting chamber, and your microscope has already been calibrated, there is no need to recalibrate. You may use the calibration factors (F) previously established for your microscope to determine sperm concentration (C) using the formula shown below (C = N x F). A good quality laboratory microscope is recommended for semen analysis. Phase contrast optics with an objective magnification of 10X to 40X are preferred for an easy visualization of the sperm cells. The Standard Count contains no counting grid, so it is necessary to use an eyepiece reticle to define the area being counted. This is best accomplished with a 10 X 10 net pattern that projects 100 boxes over the viewing field. We can provide a reticle compatible with virtually all major brands of microscopes. Calculate the Sperm ConcentrationCalculate the sperm concentration by using the following formula: C = sperm concentration If the concentration permits, we recommend counting at least 100200 sperm from a field selected in the center of the Standard Count chamber. 1. Calculate (N) by dividing the total number of sperm counted by the number of boxes counted. Obviously, you must keep track of the number of boxes counted in order to perform this calculation. 2. The factor (F) is a calibration factor designed to compensate for the optical variation that is experienced from microscope to microscope, even those of the same model and manufacturer. Once a specific microscope is calibrated and the factor F is derived, you can use that value F for all samples analyzed with the same magnification on that specific microscope. where F = the calibration factor determined for each microscope, magnification, and Standard Count chamber depth. T = the chamber depth (in microns). For the Standard Count, you would use the number 20. D = the distance across a single box of the reticle (in microns) 3. Calculate (D) by using a Stage Micrometer. To achieve this:
4. Incorporate the value (D) into the formula, and derive the factor F. Remember that the value D is squared in the formula to account for width x length. Assuming that the value of D is 25 microns and that you are using a Standard Count with a 20 micron chamber depth, the factor F would then be equal to F = 80 Determining Percent MotilityCount only the motile sperm in the boxes, but record that number. Recount the same boxes and this time count only the nonmotile sperm. Add the motile and nonmotile sperm to obtain the total number of sperm counted Calculate percent motility as follows: 