Bio-Rad Rat Diabetes Assays - Manual (Page 20)

18
6. For assays other than adiponectin and adipsin, dilute plasma fourfold
(1:4) by adding 1 volume of sample to 3 volumes of Bio-Plex sample
diluent (for example: 40 μl sample + 120 μl sample diluent).
7. Assay samples immediately or aliquot into single-use tubes and store
at –70°C. Avoid repeated freeze/thaw cycles.
Serum
1. To prepare serum, allow blood to clot at room temperature for
30 to 45 min.
2. Perform centrifugation at 1,000 x g for 15 min at 4°C and transfer the
serum to a clean polypropylene tube.
3. To completely remove platelets and precipitates, centrifuge again at
10,000 x g for 10 min at 4°C. Alternatively, carefully filter the samples
with a 0.8/0.2 μm dual filter to prevent instrument clogging.
4. Dilute and handle samples as described in steps 6 and 7 above.
Tissue 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%.
This is done 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. If high levels of analyte are expected, samples can be further diluted
in culture media. Supplement serum-free media with 0.5% BSA final.
4. Assay samples immediately or aliquot and store at –70°C.