High Select™ Top14 Abundant Protein Depletion Mini Spin Columns, 24 Columns - FAQs

Can I use the High Select Top14 Abundant Protein Depletion Midi Spin Columns to deplete proteins from rodent samples?

The High Select Top14 Abundant Protein Depletion Midi Spin Columns have only been validated for human samples. We do not recommend using them for protein depletion in rodent samples. For protein depletion in rodent samples, we recommend the following multi-species depletion resins:

• CaptureSelect MultiSpecies Albumin Depletion Product (Cat. No. 191085305)
• Pierce Albumin Serum Depletion Kits (Cat. No. 89875)


Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.

After depletion of abundant proteins from my sample, I still see albumin and other abundant proteins after LC-MS analysis. Did the depletion work?

Depletion does not remove all abundant proteins from the sample. Even with 99% depletion of a specific plasma protein, highly abundant proteins will still be the most abundant proteins in the sample detected by MS analysis. However, depletion significantly decreases the most abundant plasma proteins enabling identification of lower abundant proteins.

Find additional tips, troubleshooting help, and resources within our Mass Spectrometry Support Center.

How do I proceed to downstream mass spec sample preparation after abundant plasma protein depletion?

After the depletion, samples are significantly diluted in buffer. Depleted samples should be concentrated by speed vac drying, solvent precipitation, or diafiltration. Samples dried using a speedvac are directly compatible with the EasyPep kit chemistry after resuspension using the Lysis Solution.

Find additional tips, troubleshooting help, and resources within our Mass Spectrometry Support Center.

What is the maximum loading capacity for the High Select Depletion columns?

Mini columns can deplete up to 10 µL (600 µg) of plasma or serum; midi columns can deplete up to 100 µL (6000 µg) of plasma or serum.

Find additional tips, troubleshooting help, and resources within our Mass Spectrometry Support Center.

What is the percentage of albumin that is removed using abundant protein depletion columns?

More than 99% of albumin can be depleted when columns use the recommended sample to resin ratio.

Find additional tips, troubleshooting help, and resources within our Mass Spectrometry Support Center.

How do I assess efficiency and reproducibility of abundant protein depletion in plasma samples?

ELISA and mass spectrometry analysis are often used for evaluation of depletion performance. Protein yields after depletion can be measured using the BCA assay.

Find additional tips, troubleshooting help, and resources within our Mass Spectrometry Support Center.

I have a lot of plasma samples to prepare for mass spectrometry analysis. Is there a way to perform abundant protein depletion in a 96-well plate?

The bulk Top14 depletion resin is compatible with a 96-well filter plate such as the Agilent filter microplate (Cat. No. 200957-100). Add 600 µL of resin slurry (50%) to each of the wells in the 96-well filter plate, add 10-20 µL of sample, and incubate with gentle shaking. Depleted samples can be collected in a new 96-well polypropylene plate by centrifugation for 2 mins at 1,000 x g or by using a vacuum manifold.

Find additional tips, troubleshooting help, and resources within our Mass Spectrometry Support Center.

Do you offer a mass spec sample prep method for low abundant proteins in samples?

High Select Top14 Abundant Protein Depletion Mini Spin columns (Cat. Nos. A36369, A36370) can be used to deplete top 14 abundant plasma/serum proteins. The depletion of highly abundant proteins from the sample enables the detection and identification of low abundant proteins. The depleted samples can be processed with EasyPep MS sample prep kit to generate digested plasma/serum samples. The clean digested samples can be processed by nanoLC-MS/MS analysis for discovery experiments or nanoLC-PRM/MS analysis for targeted experiments.

Find additional tips, troubleshooting help, and resources within our Mass Spectrometry Support Center.

What formats of the High Select Top 14 Abundant Protein Depletion Resin do you offer?

The High Select Top 14 Abundant Protein Depletion Resin is offered in 3 formats:

a) Resin alone: High Select Top 14 Abundant Protein Depletion Resin (Cat. No. A36372)
b) High Select Top 14 Abundant Protein Depletion Mini Spin Columns (Cat. Nos. A36369, A36370) that can process a maximum of 10 µL of human serum or plasma
c) High Select Top 14 Abundant Protein Depletion Midi Spin Columns (Cat. No. A36371) that can process a maximum of 100 µL of human serum or plasma. For serum samples containing abnormally high amounts of albumin or IgG, the sample load may need to be reduced.

Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.

Which proteins are removed by High Select Top 14 Abundant Protein Depletion Resin?

The High Select Top14 Abundant Protein Depletion Resin uses immobilized antibodies to remove human serum albumin (HSA), albumin, IgG, IgA, IgM, IgD, IgE, kappa and lambda light chains, alpha-1-acidglycoprotein, alpha-1-antitrypsin, alpha-2-macroglobulin, apolipoprotein A1, fibrinogen, haptoglobin, and transferrin from serum, plasma, or spinal fluids.

Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.