Tissue-type plasminogen activator (tPA) is a 68 kDa serine protease that is synthesized by endothelial cells in normal blood vessels and displays relatively high affinity for fibrin, suggesting that it functions predominately in physiological thrombolysis in vivo. High level of tPA is a good prognostic marker for breast cancer. tPA may minimize the formation of metastasis by preventing tumor cell adherence at sites of trauma. On the other hand, gastrointestinal cancer is accompanied by a decrease in tPA.

Tissue-type Plasminogen Activator (tPA) Activity Assay Kit provides a simple, reproducible, and sensitive tool for measuring tPA activity in cell lysates, serum/plasma and other biological samples. This assay is based on spectrophotometric detection of the chromophore p-nitroaniline (pNA) after cleavage from the labeled substrate. Light emission from pNA can be quantified using a spectrophotometer or a microtiter plate reader at 405 nm. The change in absorbance of the pNA is directly proportional to tPA activity in the sample.

• Microplate reader capable of measuring absorbance at 405 nm
• Precision pipettes to deliver 2 µL to 1 mL volumes
• Tubes to prepare sample dilutions
• Incubator at 37°C
• 15 mL conical tubes
• MilliQ water or another type of double-distilled water (ddH₂O)

1. Add 50 µL of each sample, positive control, and substrate background to the 96-well microtiter plate.
2. Add 100 µL of each standard to the 96-well microtiter plate.
3. Initiate the reaction by adding 50 µL of tPA Substrate solution to each sample, positive control, and substrate background well. Mix thoroughly.
4. Measure absorbance signal: For kinetic reading, immediately start measuring absorbance signal at 405 nm continuously at 37°C and record data every 1 min. for 10 mins.