Bio-Rad Human MMP and TIMP Assays - Manual (Page 17)

3. Perform centrifugation at 1,000 x g for 15 min at 4°C and transfer
the serum or plasma to a clean polypropylene tube.
4. To completely remove platelets and precipitates, centrifuge again at
10,000 x g for 10 min at 4°C. Alternatively, filter the samples with a
0.8/0.2 μm dual filter to prevent clogging.
5. Dilute 5 µl sample with 245 µl diluent HD for a 1:50 dilution.
Vortex for 5 sec.
6. Assay samples immediately or aliquot into single-use tubes and
store at –70°C. Avoid repeated freeze-thaw cycles.
Cell Culture Supernatant
1. Collect supernatants and centrifuge at 1,000 x g for 15 min at 4°C.
For cell lines cultured in serum-free culture media, collect samples
and add BSA as a carrier protein to a final concentration of 0.5% to
stabilize protein analytes and to prevent adsorption to labware.
2. Transfer to a clean polypropylene tube. If cellular debris or precipitates
are present, centrifuge again at 10,000 x g for 10 min at 4°C.
3. Reconstitute and dilute the standard in the same medium or
matrix in which cells are prepared. Be sure to include all medium
components (such as FBS) as appropriate. To minimize error due
to lot-to-lot variation of culture media, use the same lot of culture
medium that was used to prepare the cells.
4. Assay immediately or store samples in single-use aliquots at –70°C.
Avoid repeated freeze-thaw cycles.
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