Recording human sperm- Calculate Hyperactivation

[natalia oscoz]

Hi all,
Continuing with my question from a few months ago. I have two questions:
The first, I use the Pylon software to record human spermatozoa, I need to study the motility, but I can't get a good recording setting, the videos look faster or slower than the original sample. Does anybody have any suggestions?
I use a Basler camera that records 200 fps and 10X negative phase contrast objective

The second question is how do I obtain the human sperm hyperactivation parameter with the values obtained with the open casa module? I understand that this parameter must be calculated.

thank you so much

Natalia.

[Felipe Martínez Pastor]

Hi Natalia,
Could you share your camera's model? Are you using all the sensor area
or a smaller size? (often, you only can achieve top speed by reducing
the frame size).
In any case, if you are sure that you are recording the correct number
of frames at the specified time (e.g., 200 frames in 1 second), then the
video should be OK for OpenCASA, even if it looks faster or slower in
VLC or other video software.
About the hyperactivation, you could get that parameter by processing
the data according to the criteria of choice (more efficient bur
requiring some programming knowledge: R or Python; easier but
time-consuming: spreadsheets and the like).
Best regards,
Felipe


El 8/3/23 a las 15:08, natalia oscoz escribió:

[Will Skinner]

Hi Natalia,

In humans, the hyperactivity cutoff values should be VCL ≥150 μm/s AND LIN ≤50% AND ALH ≥7.0 μm. Note, however, that ALH is dependent on VAP, which is dependent on choosing a reasonable window setting in OpenCASA, I believe.

There is an alternate method which does not rely on correct window settings – if OpenCASA can calculate the fractal dimension D, you could try VCL ≥150 μm s−1 AND D ≥1.20 (but in my hands on an IVOS II CASA machine I found that this measure identified about half as many cells as hyperactivated, as compared to the first metric. 

This method is detailed by Mortimer & Mortimer in the proceedings of the most recent International Symposium on Spermatology, among other places.  I

Mortimer, D., Mortimer, S.T. (2021). Routine Application of CASA in Human Clinical Andrology and ART Laboratories. In: Björndahl, L., Flanagan, J., Holmberg, R., Kvist, U. (eds) XIIIth International Symposium on Spermatology. Springer, Cham. https://doi.org/10.1007/978-3-030-66292-9_26

Good luck,

~Will

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Will Skinner

BA, Biology, Harvard '09

PhD Candidate, Endocrinology, Lishko Lab, UC Berkeley

National Science Foundation Graduate Research Fellow, 2018

Center for Emerging and Neglected Diseases Fellow, 2021

Phone: (518) 396-9892