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Pierce Mass Spec Sample Prep Kit for Cultured Cells


Optimized isolation and digestion of protein samples for mass spectrometry analysis.

Mass Spec Sample Prep Kit for Cultured Cells

The Thermo Scientific Pierce Mass Spec Sample Prep Kit for Cultured Cells is an easy-to-use, comprehensive kit for preparation of clean peptide mixtures from cultured cells for mass spectrometry (MS) analysis.

The Pierce Mass Spec Sample Prep Kit for Cultured Cells contains all the necessary reagents and enzymes to prepare up to 20 samples (1 million cells each) for MS analysis. The simple and robust workflow uses a lysis protocol that generates approximately 100μg of protein per sample and includes a control digestion indicator to monitor the efficiency of the two-step enzymatic digestion protocol. The kit procedure, which includes a Digestion Indicator as an internal protein/peptide control, produces consistently and reproducibly clean peptide mixtures for protein identification.

Highlights:

  • Complete – includes all reagents, a digestion indicator control, proteases and an optimized protocol needed to process up to 20 samples
  • Simple – user friendly kit can provide reproducible results even for non-expert MS analysts
  • Flexible – can be adapted to handle sample sizes between 10 to 200μg
  • High yield – total protein yield from 1 million cells is greater than 100μg
  • Optimized – cysteine reduction and alkylation are 100% with less than 1% over alkylation of non-cysteine residues
  • Efficient - percentage of missed cleavages is less than 10%
  • Compatible – final preparation is ready for direct MS analysis and other downstream applications, including mass-tag labeling
Video of the Pierce Mass Spec Sample Prep Kit in action!

Includes:

Kit contains proprietary lysis and digestion buffers, digestion indicator, DTT, iodoacetamide, Lys-C, trypsin

Product Details:

Sample prep for mass spectrometry (MS, mass spec) remains one of the largest bottlenecks associated with MS analysis, and consistent and reproducible sample preparation can make the difference between a successful analysis and a failed outcome. Current sample prep protocols are primarily homebrew and can be highly variable, making data analysis and interpretation difficult. The Mass Spec Sample Prep Kit for Cultured Cells provides researchers with all the necessary tools to generate consistent and reproducible protein digests that are directly compatible with LC-MS workflows. The digests do not require further processing, such as C18 clean up or detergent removal.

The Pierce Digestion Indicator, which can be purchased separately, is a non-mammalian recombinant protein (26kDa) with five signature peptides for use in determining the digestion efficiency and reproducibility across multiple samples. The protein sequence and recommended peptides to monitor across samples are supplied with the product instructions.

Pierce Mass Spec Sample Prep Kit for Cultured Cells protocol summary.
Protocol summary for the Thermo Scientific Pierce Mass Spec Sample Prep Kit for Cultured Cells. All reagent and enzymes are supplied in the kit, except trifluoroacetic acid (TFA), phosphate-buffered saline (PBS), and acetone. Pierce Universal Nuclease for Cell Lysis (Part No. 88700) can be used as an alternative to sonication for cell lysis.

 

Comparison of protein yields by four mass spectrometry sample prep methods.

Comparison of protein yields by four mass spec sample prep lysis methods. From one culture of HeLa S3 cells, duplicate pellets containing 2 x 10^6 cells were resuspended and lysed using 0.2mL of the respective buffers and protocol of each method (all include a sonication step); then protein concentrations and yields were determined.

  • FASP: 0.1M Tris-HCl, 4% SDS, 0.1M DTT, pH 7.6
  • AmBic-SDS: 0.05M ammonium bicarbonate, 0.1% SDS, pH 8.0
  • Urea: 0.1M Tris-HCl, 8M urea, pH 8.5
  • Kit Buffer: Lysis Buffer from the Pierce Kit (Part No. 84840)

 

Consistent, clean and reproducible LC-MS/MS results of three biological replicates. HeLa cell lysate (200μg in 200μL of Lysis Buffer and spiked with 2μg Digestion Indicator) was processed by the Thermo Scientific Pierce Mass Spec Sample Prep Kit for Culture Cells and then analyzed by mass spectrometry.
  Sample 1 Sample 2 Sample 3
Number of Proteins 3382 3228 3376
Number of Unique Peptides 16333 15939 17048
Missed Cleavages (%) <10 <10 <10
Disulfide Bond Reduction (%) 100 100 100
Cysteine Alkylation (%) 100 100 100
Over Akylation (%) 0.1 0.3 0.9
Digestion Indicator Protein
Sequence Coverage (%)
62.50 62.93 65.09

 

More Product Data

References:

  1. Wisniewski, JR., et al. (2009). Universal sample preparation method for proteome analysis. Nat. Methods. 6:359-60.
  2. Bereman, MS., Egertson, JD., MacCoss, MJ. (2011). Comparison between procedures using sodium dodecyl sulfate for shotgun proteomic analyses of complex samples. Proteomics 11:2931-5.

Related Resources:

Mass Spectrometry Sample Preparation Handbook
Review of mass spectrometry

Related Products:

In-Solution Tryptic Digestion Kit
In-Gel Tryptic Digestion Kit
Pierce C-18 Spin Columns
DTT (dithiothreitol)
Proteases for Mass Spectrometry


Ordering Information
 
Product # Description Pkg. Size Instructions MSDS CofA Price
84840 Mass Spec Sample Prep Kit for Cultured Cells
Formulation: Multi-component kit
Sufficient For: 20 protease digestions and preparations from 100μg of cell lysate

Kit Contents:
Cell Lysis Buffer, 5mL
Digestion Buffer, 5mL
DTT, No-Weigh Format, 24 tubes
IAA, No-Weigh Format, 24 tubes
Trypsin Storage Solution, 250μL
Pierce Digestion Indicator, 10μg
Lys-C Protease, MS Grade, 20μg
Trypsin Protease, MS Grade, 2 x 20μg
20-rxn kit Product Instructions for product #84840 Mass Spec Sample Prep Kit for Cultured Cells MSDS for product #84840 Mass Spec Sample Prep Kit for Cultured Cells Certificate of Analysis for product #84840 Mass Spec Sample Prep Kit for Cultured Cells $405.00

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84841 Digestion Indicator for Mass Spectrometry
Formulation: Non-mammalian recombinant protein (26kDa), frozen liquid
Sufficient For: Production of five signature peptides upon digestion for mass spectrometry
10µg MSDS for product #84841 Digestion Indicator for Mass Spectrometry Certificate of Analysis for product #84841 Digestion Indicator for Mass Spectrometry $154.00

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Instructions | MSDS | CofA
Product Instructions | MSDS | CofA  

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