Alpha-1-Microglobulin (a1M) is a small protein belonging to the superfamily of lipocalins, which is present in various body fluids such as plasma and cerebrospinal fluid. it has a role in the transportation of molecules like metals and serves as a marker for certain diseases, including kidney and liver disorders.The activity of recombinant human A1M was measured by its ability to inhibit trypsin cleavage of a fluorogenic peptide substrate MCA-Arg-Pro-Lys-Pro-Val-Glu-NVAL-Trp-Arg-Lys(DNP)-NH2 in the assay buffer 50 mM Tris, 10 mM CaCl2, 150 mM NaCl, 0.05% (w/v) Brij-35, pH 7.5. Trypsin was diluted to 50 ug/ml in the assay buffer and 25 ul different concentrations of recombinant human a1M (MW: 64 KD) was incubated with 25ul diluted trypsin at 37 ℃ for 15?minutes.Loading 50?μL of the incubated mixtures which were diluted five-fold in assay buffer into empty wells of a plate, and start the reaction by adding 50?μL of 20?μM substrate.nclude a substrate blank containing 50?μL of assay buffer and 50?μL of 20?μM substrate.Then read at excitiation and emission wavelengths of 320 nm and 405 nm, respectively, in kinetic mode for 5 minutes.The result was shown in Figure 1 and it was obvious that recombinant human a1M significantly decreased trypsin activity. The inhibition IC50 was <58 nM.