Biofluid isolation

We follow the recommendations provided by National Cancer Institute's Early Detection Research Network:

Tuck MK, Chan DW, Chia D, Godwin AK, Grizzle WE, Krueger KE, Rom W, Sanda M, Sorbara L, Stass S, Wang W, Brenner DE. Standard operating procedures for serum and plasma collection: early detection research network consensus statement standard operating procedure integration working group. J Proteome Res. 2009 Jan;8(1):113-7. doi: 10.1021/pr800545q. PMID: 19072545; PMCID: PMC2655764.

Plasma Isolation

1. After blood collection into EDTA vacutainer, store upright until ready for centrifugation.
2. Centrifuge within 4 hours of collection, but sooner is better.
3. Spin tubes down for 10-20 minutes at 1,100 rcf in a room temperature microcentrifuge.
1. If samples slated for EV enrichment by Mag-Net, we strongly recommend avoidinghigh speed spins that are suggested in other protocols to reduce platelet content. This also reduces EV content.
4. EDTA tubes should be carefully removed from the centrifuge.
5. The upper layer in the tube is plasma, and should be removed carefully with a pipette (Do not pour).
1. NOTE: Avoid disturbing the buffy coat. Leave a small amount of plasma behind.
6. Place separated plasma on wet ice.
1. If multiple pipette transfers are needed, pool them together in a tube and mix by hand.
1. NOTE: EVs can be damaged by vortexing - we do not recommend this.
7. For best results, aliquot plasma upon collection in smaller volumes depending on planned experiments to reduce freeze-thaw cycles later.
8. Store at -80C, and ship samples on dry ice.

Serum Isolation

1. After blood collection into red top (silicon coated, no additives, no SST/serum separator tubes) vacutainer, store upright for 30-60 minutes to allow clot to form.
2. Centrifuge at the end of the clotting time (30-60 minutes) in a horizontal rotor centrifuge for 20 minutes at 1,100 rcf in a room temperature microcentrifuge.
1. If centrifugation cannot be performed at the conclusion of clotting, the tubes should be refrigerated for no more than 4 hours.
3. Tubes should be carefully removed from the centrifuge.
1. The upper layer in the tube is serum, and should be removed carefully with a pipette (Do not pour).
2. NOTE: Avoid disturbing the red blood cell layer. Leave a small amount of serum behind.
4. If multiple tubes or pipette transfers from the same individual are needed, pool them together in a tube and mix by hand.
1. NOTE: EVs can be damaged by vortexing - we do not recommend this.
5. For best results, aliquot serum upon collection in smaller volumes depending on planned experiments to reduce freeze-thaw cycles later.
6. Store at -80C, and ship samples on dry ice.

Experimental design to reduce batch effects

We recommend reviewing the following publications to begin intergrating steps from the beginning of your experiment to reduce batch effects:

It is crucial to ensure that not all samples representative of an exerimental group are collected or processed together. For example - if all control samples are collected one day, and all treated samples are collected on another day, it is very difficult to deduce whether an associated phenotype measured is not effected by this. Collecting half of the control samples and half of the treated samples on one day, and then the other half of both groups on another day at least reduces the likelihood of a batch effect strongly skewing the entire study.

We use the metadata you share to ensure that samples are randomized such that no experimental groups are over-represented during a single sample preparation session.

Sample shipment

 Please contact us and confirm your plans before dropping samples in the mail to reduce risk of sample loss! Refer to our Shipping page for more details.