Hi, I'm having trouble creating a mass calibration when using slides coated in poly-L-lysine (for r-loop + protein interaction work). BSA, beta amylase, and thyroglobulin are sticking to the slides and I've had some success with beta gal. Are there any protein recommendations?
Is there any documentation for this?
I read on the vial that the calibrant is 500X and a volume of 50 uL. Can I prepare 20 uL aliquots at 2X and store them at -20 to avoid repeated freeze/thaw cycles?
Any information is welcome.
Thanks,
Ryan
Hi Ryan,
We have not tested “20 uL aliquots at 2X” and hence unable to comment on the product stability at such dilutions. But what we can say is for best results, aliquot 10 x 5 µL upon first thaw and store to avoid repeated freeze/thaw cycles.
Best,
Team Refeyn
Hi, what affects the % error for the massference P2 calibrants? I tried doing 2 calibrants for a slide and the % error is around 4%.
i would appreciate a response. Thank you!
Hello,
The mass precision (CoV) of the SamuxMP is 2%, meaning that is 4% difference in results is possible, but it is rare that it would be this large. There are a couple of things which can affect this (and the factors that can cause mass deviations for MFP2 are the same factors that can cause deviations for any other measurement):
Temperature: The temperature inside the system effects the contrasts and therefore the mass measured by the mass photometer. We would therefore recommend calibrating regularly (at least every 2 hours) and ensuring your system has been turned on for around an hour before use so it can reach thermal equilibrium.
Bubbles in immersion oil: Bubbles in the immersion oil can cause shifts in the measured mass. The latest versions of the software should detect these bubbles, but you can also look for breaks in the focus ring which can indicate a bubble in the oil. If you find one, it can be moved by removing the magnets and lifting the slide at one side enough to break the connection between the oil drop and the slide.
I hope this helps but please feel free to reach out again if you have any further questions.
Best regards,
Refeyn
Hi Nicole,
Can I ask what you mean by sticking to the slides? We measure the particle signals as they bind to the slide. If you're not see individual particles during the recording, the concentration might be off. Try using 5-10 nM of protein for measurement
It might be easier to look at the data and provide better troubleshooting help. Feel free to email us at support@refeyn.com or to me directly at amy.chau@refeyn.com. We can discuss more in detail via email or call.
Amy Chau
Sr. Field Applications Scientist