Table 2.
Surface mix
| Marker and Clone | Fluorophore | Volume (for 100 μL cell suspension per sample) | Dilution |
|---|---|---|---|
| CD3 (SK7) | APC-H7 | 0.6 μL | 1:167 |
| CD4 (L200) | BV711 | 0.3 μL | 1:333 |
| CD8 (SK1) | BUV805 | 0.6 μL | 1:167 |
| CD45RO (UCHL1) | PE-Cy™7 | 2.5 μL | 1:40 |
| CD19 (SJ25C1) | BV786 | 0.6 μL | 1:167 |
| CD56 (NCAM16.2) | BUV395 | 0.6 μL | 1:167 |
| CD27 (M-T271) | Alexa Fluor 700 | 5 μL | 1:20 |
| CD21 (B-ly4) | BV650 | 1.25 μL | 1:80 |
| Brilliant Stain Buffer | N/A | 88.55 μL | NA |
| Total | 100 μL | NA | |
The preparations provided here is for 1 tube. The volume has to be multiplied by the number of tubes: for one sample you would have to make the surface mix times 3 (fully stained untreated sample, FMX untreated sample, fully stained treated sample). In case of a higher number of samples, it is suggested to prepare a master mix with at least 10% extra volume (e.g., if 10 tubes are required, calculate on the basis of 11), so that even the last tube is filled properly avoiding the effect of sample loss when dosing.
The mAbs have to be titrated for identifying the best concentration that allows to distinguish the populations of interest.