Mouse sperm

[Balma García]

Hello everyone, 

I would like to start using the CASA system with mouse sperm. I have installed the program already, but I have some basic questions, if you could help me...

1. To capture images should I use a 10x objective or can it be 20x?

2. Can I modify the values that the ImageJ program extracts? When I try to do something about the image the program doesn't detect it.

3. Do I have to set some default value? How can I do it?

Any advice before starting? Any idea or tip will be welcome.

Thank you

[Erick José Ramo Da Silva]

Dear Balma,

We have been working with Carlos to standardize OpenCasa for mouse spermatozoa.

So far the results are promising, but the fact their sickle-shaped head makes it a little difficult to build proper tracks.

In response to your questions:

1. To capture images should I use a 10x objective or can it be 20x?  

Typically evaluation of rodent (mouse and rat) sperm motility by CASA should be performed in a 4X objective. They are larger cells (in length) than human and bull spermatozoa, for instance.

For a very good and detailed protocol on CASA analysis for mouse spermatozoa, refer to Goodson et al. Biol Reprod, v. 84, n. 6, p. 1207-15, 2011. doi. 10.1095/biolreprod.110.088989.

2. Can I modify the values that the ImageJ program extracts? When I try to do something about the image the program doesn't detect it.  

I don't know exactly what you mean by "modify the values", but you can export all the data to an excel spreadsheet, which is much easier to work with.    

3. Do I have to set some default value? How can I do it?

Yes, you will need to make some changes on the settings since mouse spermatozoa requires specific parameters for proper detection in OpenCasa.

We have performed several tests using OpenCasa 1.0 (we have not tested yet in OpenCasa 2.0). Find attached our settings, which were defined for videos captured using a HT Ceros 2 system and a 4X objective. 

They are rather experimental and seem to detect progressive motile tracks nicely, but are not good if you want to evaluate hyperactive tracks. So additional fine tuning on the settings are yet required.

I hope it helps. 

Feel free to share your thoughts on our settings.

Kind regards,

Erick

Erick J. R. Silva, Ph.D.

Assistant Professor

Department of Biophysics and Pharmacology

Institute of Biosciences of Botucatu

Universidade Estadual Paulista "Júlio de Mesquita Filho" (UNESP)

Rua Prof. Dr. Antonio Celso W. Zanin, S/N

Botucatu-SP, Brasil, 18618-689 

Phone: +55 14 3880-0223
E-mail: ejr.s...@unesp.br // ersi...@gmail.com

Researcher ID: D-2976-2009

Scopus ID: 36176142500

ORCID: https://orcid.org/0000-0002-9330-8658

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[Balma García]

Dear Erik,

thank you for your answer. I hope that together we can standardize the parameters.

I have installed version 1 but it won't let me open the videos I have, that's why I'm trying with version 2. I've been able to change the settings and add the ones you gave me.

Before continuing I would like to ask some basics about the videos. I know the format has to be .avi. But what about frames and pixels? Are there any requirements? Maybe this is why I can't open the videos with v.1.

I am testing the ImageJ with the Test videos, and some sperm does not detect by the program. I'm attaching images. To analyze them I have put the appropriate parameters for the test.

Do you have any suggestions ? I start from 0, so any suggestion is welcome

Best,

Balma.