How do we perform TGFβ-1 lab test?
Protocols we follow to perform TGFβ-1 tests
We use Luminex, ELISA, and MSD assays to detect TGFβ-1 in serum, plasma, and tissue/cell culture samples. Besides the criteria listed in the protocol, we also test dilution linearity, cross-reactivity, kit stability, and sample detectability.
Following is an elaborated Luminex protocol.
We begin with 200 μl of assay buffer in each well and shake it for 10 mins at RT. We decant the assay buffer and residues and introduce 25 μl of the following ingredients in appropriate wells: Standards, control or treated samples, followed by assay buffer, matrix solution, , and beads and incubate it at RT for 2hrs or overnight at 4 °C. After incubation, plates are incubated for 2 mins on magnetic plate, following which well contents are removed and washed with a 200 μl wash buffer. Once the beads capture a sample analyte, we incubate them with a biotinylated detection antibody. We then complete the reaction by incubating with 25 μl Streptavidin-Phycoerythrin, a reporter molecule in each well for 30 mins at RT. Post-incubation, we remove the well contents and wash them with a 200 μl wash buffer and add 100 μl of drive or sheath fluid. We read the assay on an appropriate Luminex platform.
The transforming growth factor beta 1 test or assay identifies each microsphere through fluorescent reporter signals. We can perform this assay in a single day or over two days with overnight incubation and it requires 25 μl of 1:30 dilution serum/plasma post-treatment. The assay has sensitivity between 8-12 pg/ml and an accuracy of 62-128 %.