Target cell killing assay: Cr51 method
Target cell killing assay: Cr51 method
Detection of killing activity of effector cells
Principle
The target cell is labeled with Na2 51 CrO 4, and if the effector cell to be examined can kill the target cell, then 51 Cr (chromium) is released from the target cell. The released51 Cr radioactivity was measured by γ-counter. The more the target cells were lysed and destroyed, the more51 Cr was released, and the higher the radioactivity of the supernatant was, and the killing activity of the effector cells to be examined could be calculated by applying the formula.
Appliance
Oncology experiments
Operation method
Target cell killing assay: Cr51 method
Materials and Instruments
[Materials] Na2 51 CrO 4, effector cells; target cells; Move (CTL killing of tumor cells as an example):1、Effector cellsSeparated peripheral blood T cells, cultured for 7d, according to the ratio of DC:T = 1:10, add their own cell-sensitized DC, cell line-sensitized DC and unsensitized DC, and continue to culture for 3-4d, then collect the T cells, which is the effector cell CTL.2. Target cells1×106 Panc1 and pancreatic cancer cells (S1-S6) isolated from 6 tumor tissues were collected and resuspended in 0.5 mL of complete culture medium without sodium bicarbonate. Add 3.7 MBq 51Cr sodium chromate, label for 1 h at 37℃ in a water bath, wash the cells three times, and adjust the cell concentration to 5×104/mL.3. Cytotoxic effectAdd 100 μL of target cells into each well of 96-well plate. Add 100 μL of different effector cells into each well at the ratio of effector cells:target cells=50:1. For each group, there were 3 duplicate wells, and only 100 μL of complete culture medium was added to the negative control wells (spontaneous release), and 100 μL of 1% NP40 (v/v) was added to the positive control wells (maximal release). The 96-well plates were incubated at 37℃ for 4 h. After centrifugation of the plates at 1000 g for 10 min, 100 μL of supernatant was aspirated from each well and the activity per minute (cpm) was measured on a liquid flash counter. Caveat Cell killing activity was calculated according to the following formula: CTL activity (%) = [(experimental group cpm - spontaneous release cpm)/(maximum release cpm - spontaneous release cpm)] × 100%.Statistical processing was performed using SPSS statistical software, and all variables were expressed as mean±SD. Comparison of the significance of differences between variables was performed by analysis of variance (Student-Newman-Keuls method). For more product details, please visit Aladdin Scientific website.