Dear Navid, if all the equipment are precisely adjusted (the scale, pipettes and the pH meter) so try to adjust cautiously by microliters of HCl and/or NaOH (1N) according to the desired pH, and then wait for mixing by stirring then re-measure by the pH meter. Be careful, addition of large amount of either acids or bases harms the sperms and oocytes.

Dear Islam, thank you so much for the answer. Do you have any idea about HCl molarity for this adjustment?

(1N) means Normality unit equals 1 or a normal HCl solution; you can calculate it online 

http://www.sigmaaldrich.com/chemistry/stockroom-reagents/learning-center/technical-library/molarity-calculator.html

Dear Islam, thank you so much.

Dr. Navid... I prepare Sp-TALP with HEPES and rarely I have problems with pH... I use Sp-TALP with HEPES to flow cytometric procedures and induction of acrosome reaction in stallion and bull sperm... This could be another option for adjust pH in culture media

Before making the final dillution, adjust the pH,  check the electrode on the pH meter, and calibrate properly on the day of use, sometimes this can be a factor. We adjust most of the media a room temp; and after preparing the media we take an aliquot, place it in the respective incubator and check the pH in the incubator after a couple of hours of aquilibration,  remember that CO2 will low the pH of the media in the incubator. 

To adjust pH we maintain a 1M and 7M solutions, and use them depending in the need. 

Dear Navid,

I understand why you decided to do in-house media. If I may bring this up for your consideration. One question you would have to ask to yourself is what is less expensive: keep buying commercial media or mastering the in-house preparation. Eventually you will reach proficiency but you need to invest time and money if you are able to afford it.

 Answering your technical question about pH

For storage we would want to adjust the pH. On the other hand, once the media reaches set temperature and/or calibrated to CO2 conditions pH will adjust accordingly. 

For holding media (atmospheric conditions), I would agree with everyone just make sure there is a zwitterionic salt included (HEPES is one example). I usually prepare 100 mL of COC holding media (TCM-199 HEPES, FCS HI, Gent, Na-Pyr and Heparin) and before filtering I just need to add 100uL of NaOH to reach a pH of 7.3-7.4.

As you may know, all your media exposed to CO2 conditions would need to have HCO3-.  I would agree with Sofia, the more accurate way of reading your pH under your settings is by measuring pH once the medium has been calibrated in the incubator, although I would like to use larger amounts  and a period of minimum 4-5 hrs. Then, the less expensive way is to quickly remove your tube with media, insert the pH probe and get your reading asap. If you do not feel comfortable with that there are more expensive probes that continuously monitor pH.

http://www.origio.com/products/scanlab%20equipment/safe%20sens.aspx

Lastly you would just need to adjust your CO2 levels, this is why sometimes you see in the articles 5% or 6% CO2, pretty standard.

For all your culture media not containing zwitterionic salts try to minimally expose your droplets or 4-well dishes to atmospheric conditions at any checkpoint. Light mineral oil overlay helps to minimize pH fluctuations whenever you need to take the dish out of the incubator. I agree with Islam make sure your equipment is properly calibrated (this includes incubators), very critical.

Best wishes,

Dear Sofia,

thank you so much for the critical points.

Regards,

Navid

Dear Victor,

Thank you so much for grate and complete answer.

Have a nice day!

Navid

Dear Navid,

If you want to prepare SP-TALP you have to add HEPES (10-20 mM) as a buffer.

We use 100 mM HCl and NaOH for adjustment of pH in our lab. You can add phenol red to your solution, thanks to phenol red it is possible screen pH shifts in media. You can add phenol red at 0.03 mM in your solution without any harmful effects on cells. I agree with Dr. Victor who notified about use of NaHCO3 buffer systems with CO2 incubator and use of HEPES buffer in holding and manipulation media which used in ambient conditions. The solution’s pH also change with temperature.

With Best Regards

Dear Shahram,

Thanks for the kindness and sharing your valuable experience.

regards

Hi Navid, we usually do not adjust the pH of the medium. At preparing sodium bicarbonate is used so that the pH is adjusted alone incubated it about 4-6 hrs before using it, in a tube with a loose lid that will take effect CO 2 from the stove, which usually is maintained at 5%. You can check de pH after 6 hrs of incubation.

The same goes for the Fert-TALP

Cheers and hope to help you