Alanine aminotransferase activity assay kit

Three groups of 7-week-old female C57BL/6 mice were repetitively injected with PBS, 5 mg per kg of dNP2-dTomato or dNP2-ctCTLA-4, respectively, every other day for 14 days. The weight changes of the mice were daily monitored. At day 15, the mice were sacrificed and the morphologies of the spleen, liver and brain were carefully observed. The cytotoxicity of each protein against splenocytes and thymocytes was analysed using an Annexin V and 7-AAD staining kit (BD bioscience). The percentages of naive CD4 T cells in lymph nodes and spleen were analysed with isolated lymphocytes from each tissue after staining with 1/800 diluted anti-mouse CD4-PerCP-Cy5.5 (#45-0042), anti-mouse CD62L-FITC (#11-0621) and anti-mouse CD44-PE (#12-0441) FACS antibodies purchased from eBioscience. Liver toxicity of the proteins was analysed using an alanine aminotransferase activity assay kit (BioVision) and an aspartate aminotransferase activity assay kit (BioVision).

An alanine aminotransferase activity assay kit (BioVision, Inc.) was used to measure the activity of ALT. A mixture of L-alanine, NADH, LDH (lactate dehydrogenase) and ALT was incubated at 37 °C for 15 min, then α-ketoglutarate was added to the mixture and the absorbance at 340 nm was measured with a microplate reader.