The immunotherapy field is making great progress in aiding many disease treatments. The hope is that one day, personalized immunotherapy can be used so each patient can use their own immune system to overcome their specific disease affliction. QuickSwitch is a tool for creating custom MHC class I tetramers quickly and easily.
Have you seen comparisons of multimer technology claiming to be superior to tetramers? Be sure to ask which tetramers were used in the comparison! MBL tetramers have a clear advantage over academic tetramers and other commercial MHC multimer products, not only due to the reliability and high quality for every lot produced, but also due to the proprietary alpha-3 mutation. This mutation, engineered into the heavy chain of all of our human and macaque class I tetramers, helps decrease non-specific binding, leading to enhanced specificity. Check out the images below to see the details of this important technology!
You’re sitting at your flow cytometer, staring at your CD8 x tetramer plot in anticipation of a tiny but meaningful group of dots appearing in the upper right quadrant. You hold your breath, click “acquire,” and watch… Wait, what? What’s this ugly diagonal?? How come ALL the CD8 positives are dual positive??! Hey, this is my control mouse! That one should be completely tetramer negative! Yep, I’ve been there too. Classic case of using a CD8 antibody that doesn’t play nice with tetramers.
In the previous Tetramer Tips blog, I suggested ways for you to be a negative Control Freak. Now it’s time to think positive! A positive control for a tetramer is a sample that contains cells expressing the specific T cell receptor of interest, i.e. has the exact specificity of the tetramer. Okay, brace yourselves; I’m going to be frank here. Having a positive control for tetramer experiments is often a quest for the Holy Grail. In many (most, in fact) cases, you will not have access to a positive control, unless one of your experimental samples happens to show a positive result. In an experiment where no positive events are seen and no positive control was used, you cannot necessarily conclude that the donor/patient/mouse is negative for that T cell specificity, because, heck, maybe you got distracted while pipetting and forgot to add the tetramer to your staining cocktail! A tetramer experiment with no positive control and no positives in the experimentals is, therefore, uninterpretable. This is the hard truth we must face.
While being a Control Freak may not be the best choice for living in spiritual harmony, it is a downright asset in flow cytometry research! Controls not only help you set up an experiment to get a clear or “true” answer, they can also help you troubleshoot what may have gone wrong when your data just doesn’t look quite right, so that your next attempt will turn out better. In flow cytometry, controls are critical to help determine “real” events from artifacts. So, what should you use as controls in tetramer experiments?