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Home › Learning centers › Protein research › Mass spectrometry digestion reagents › Capturem technology for trypsin protein digestion

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Capturem 96-well plate trypsin for mass spectrometry HTP Capturem trypsin
Tech Note

Protein digestion with Capturem Trypsin

Capturem membranes are a revolutionary technology that consist of porous, high-capacity membranes that can be functionalized with not only proteases (trypsin or pepsin) for digestion but also ligands such as Protein A or G, nickel, and streptavidin for purification. We've assembled these functionalized membranes into a number of different spin-column, multiwell-plate, and large-volume formats to bring you effortless, high-quality results across your protein research workflow.

Digestion workflow  

The high local trypsin concentration in the column plus the large internal surface area across a short distance ensures almost instant binding and cleavage of proteins and antibodies at room temperature. Complete antibody digestion takes 3–6 minutes as compared to traditional methods that may require overnight digestion.

Capturem Trypsin digestion workflow

Figure 1. Capturem Trypsin digestion workflow. After initial activation of the columns with the included Capturem Trypsin Activation Buffer, the sample solution (50–800 µl) containing the native or denatured sample is passed through the spin column via two-minute centrifugation. Digestion by trypsin occurs on the column, and subsequent peptide fragments are eluted in a second two-minute centrifugation. *Denaturation/reduction/alkylation time is not included.

Data  

High well-to-well reproducibility

A well-characterized protein commonly used as a standard in peptide studies, apomyoglobin (Apo), was used to test the reproducibility of Capturem Trypsin plates. Apo was digested under native conditions using three different wells of Capturem Trypsin 96-well plates. All three digests yielded nearly identical HPLC profiles (Figure 2), thus demonstrating the reliability of the Capturem Trypsin protocol. Here we show the digestion of proteins within ~2–3 minutes using Capturem Trypsin 96-well plates.

HPLC profiles well-to-well reproducibility of apomyoglobin digests using Capturem Trypsin

Figure 2. HPLC profiles well-to-well reproducibility of apomyoglobin digests using Capturem Trypsin. Tryptic spin digests of 80 μg apomyoglobin under native conditions give reproducible HPLC profiles.

Quantitation using Capturem Trypsin

Capturem 96-well trypsin plates allow specific peptide fragments to be detected and quantified in a high-throughput manner (Figure 3) with excellent well-to-well reproducibility. We performed and analyzed the tryptic spin digest of SILuLite. The quantitative analysis showed that Capturem Trypsin digestion yielded surrogate peptides detectable at ng-input amounts (1, 10, and 50 ng).

Reliable quantitation

Figure 3. Reliable quantitation. Tryptic spin digest of SILuLite yields surrogate peptides detectable at ng-input levels (1, 10, 50 ng). AUCs of specific peptides give a linear correlation.

Chemistries & formats  

TechnologyFormatTypical yieldTimeMaximum single load volume
Capturem Trypsin/Pepsin Miniprep 25 μg IgG2 5 min3 850 μl
96-well plate1 25 μg IgG2 15 min3 1 ml
Capturem Protein A Miniprep 40 μg 5 min 850 μl
96-well plate 40 μg 15 min 1 ml
24-well plate 400 μg 15 min 4 ml
Maxiprep 1 mg 15 min 23 ml
Capturem Protein G Miniprep 60 μg 5 min 850 μl
96-well plate 60 μg 15 min 1 ml
24-well plate 600 μg 15 min 4 ml
Maxiprep 1.2 mg 15 min 23 ml
Capturem Streptavidin Miniprep 25 μg IgG4 5 min 850 μl
96-well plate 25 μg IgG4 15 min 1 ml
Capturem His Miniprep 80 μg 5 min 850 μl
96-well plate 80 μg 15 min 1 ml
24-well plate 800 μg 15 min 4 ml
Maxiprep 1.5 mg 15 min 23 ml
Large volume 8–20 mg 30 min 500 ml

1Not available for Capturem Pepsin
280 μg protein
3Excludes denaturation/reduction/alkylation time
44,000 pmol free biotin

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Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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Notice to purchaser

Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval.

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