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Pierce™ Peptide Desalting Spin Columns (Thermo Scientific™)

Thermo Scientific Pierce Peptide Desalting Spin Columns are ready-to-use centrifuge spin columns that enable efficient desalting of peptide samples following enzymatic digestion. The polymer-based hydrophobic resin contained in each column provides excellent binding and recovery characteristics for peptide samples in preparation for mass spectrometry and other methods.

Features of Pierce Peptide Desalting Spin Columns include:
Removes MS-interfering contaminants—significantly reduces signal suppression and improves sensitivity in mass spectrometry
High binding capacity—special hydrophobic polymer-based resin allows for excellent recovery of peptide loads of 5 µg to as high as 5 mg of native and TMT-labeled peptides
Easy to use—resin is provided in single-use spin column format that fits many common 2 mL tubes
Compatible—resin is validated with a variety of complex samples, including TMT-labeled peptides, and is stable under extreme pH conditions

Pierce Peptide Desalting Spin Columns provide a convenient and reproducible way to desalt and remove contaminants from peptide samples prior to analysis by mass spectrometry. Each spin column can bind from 5 µg to 5 mg of native and TMT-labeled peptides. The spin column format allows processing of multiple samples (10 to 300 µL each) in parallel in approximately 30 minutes.

Mass spectrometry is essential for studying biological compounds. However, many of the buffers and reagents used in the course of sample preparation will interfere with mass spectrometry analysis, leading to poor-quality spectra and reduced sensitivity. Pierce Peptide Desalting Spin Columns are designed to remove many of the interfering hydrophilic sample contaminants with minimal sample loss, thus enabling sensitive and comprehensive analysis of digested protein samples by mass spectrometry.

Pierce™ C18 Spin Columns (Thermo Scientific™)

Thermo Scientific Pierce C18 Spin Columns are ready-to-use centrifuge columns of porous C18 reverse-phase resin with excellent binding and recovery characteristics for peptide sample preparation for mass spectrometry and other methods.

Peptide samples can be purified and concentrated for a variety of applications using Pierce C18 Spin Columns. Each spin column contains a porous C18 reversed-phase resin with excellent binding and recovery characteristics for a wide range of peptide concentrations. The spin column format allows simultaneous processing of multiple samples (10 to 150 µL each) in approximately 30 minutes without laborious repeat pipetting or specialized equipment. Pierce C18 Spin Columns can be used effectively for processing peptides derived from 10 ng to 30 µg of protein. Sensitivity and detection limits are dependent on the downstream application.

Features of C18 Spin Columns:

Removes interfering contaminants—significantly reduces signal suppression and improves signal-to-noise ratios and sequence coverage
Simplifies optimization—processing yields high-quality spectra and is effective for a variety of reverse-phase-compatible contaminates
Robust—works with a wide variety of load volumes and concentrations; no need to reduce sample volume before application
Convenient—easy to handle and requires no special equipment to process multiple samples simultaneously (unlike tip-driven systems that require one sample to be processed at a time)
Sensitive—special C18 resin allows excellent recovery percentages, even at low (sub-picomole) sample loads

Matrix-assisted laser desorption ionization (MALDI) and electrospray ionization (ESI) mass spectrometry (MS) are essential for studying biological compounds because of the high sensitivity and mass accuracy they provide. However, many of the buffers and compounds common to biological samples interfere with both MALDI-MS and ESI-MS, resulting in reduced sensitivity and poor-quality spectra. Pierce C-18 Spin Columns remove interfering contaminants and release peptides in MS-compatible solutions, providing an effective tool that aids in the MS examination of post-translational modifications or for the identification of proteins by peptide mapping.

Pierce™ Graphite Spin Columns (Thermo Scientific™)

Thermo Scientific Pierce Graphite Spin Columns improve phosphopeptide analysis by efficiently binding hydrophilic peptides and efficiently removing urea, salts and other contaminants before mass spectrometry analysis.

Features of Graphite Spin Columns:

• Convenient spin format for parallel processing of multiple samples
• High-binding capacity with excellent recovery of up to 100 µg of hydrophilic peptides per column
• Porous graphite resin for cleaning up phosphopeptide samples before MS analysis

Pierce Graphite Spin Columns enable fast and efficient capture, concentration, desalting and elution of hydrophilic peptides in less than 10 minutes. These columns are ideal for improving mass spectrometric (MS) analyses of samples from protein digests), strong-cation exchange fractions, and enriched phosphopeptides eluted from titanium dioxide and immobilized metal affinity chromatography (IMAC) columns and tips.

The Pierce Graphite Spin Columns improve phosphopeptide analysis by efficiently binding hydrophilic peptides and efficiently removing urea, salts and other contaminants before MS analysis. The C18 resins and C18 tips that are commonly used to desalt peptides are excellent for use with hydrophobic peptides but do not efficiently capture hydrophilic peptides, like phosphopeptides, resulting in enrichment of only hydrophobic fragments. The Pierce Graphite Spin Columns address this issue and are ideal for matrix-assisted laser desorption ionization (MALDI) or nanoelectrospray ionization techniques.

Pierce™ C18 Spin Tips (Thermo Scientific™)

The Thermo Scientific Pierce C18 Spin Tips enable fast and efficient capture, concentration, desalting, and elution of up to 10 µg peptides per 20 µL sample for mass spectrometry (MS) analysis.

Pierce C18 Spin Tips are 20 µL-capacity pipette tips with accompanying adaptors for microcentrifuge sample processing. The tips contain a C18 reversed-phase sorbent that minimizes flow resistance and provides excellent binding and recovery characteristics at a wide range of peptide concentrations upstream of matrix-assisted laser desorption ionization (MALDI) or nanoelectrospray ionization techniques. Sample clean-up with C18 resin significantly improves protein analysis results by removing urea, salts, and other contaminants before mass spectrometry (MS). Each Pierce C18 Spin Tip has a 20 µL volume capacity with a 10 µg peptide-binding capacity.

Features of C18 Spin Tips:

Rapid—C18 fast-flow tips have low resistance and improved flow characteristics compared to other commercially available tips
High capacity—up to 10 µg peptide per 20 µL solution
Convenient—spin tips come with tip adaptors for easy centrifugation
Cleaner sample—device design filters out particulates that can cause autosampler and column clogging

Applications:
• Clean up of peptides after protein digestion
• Peptide sample concentration and desalting
• Sample preparation for MALDI peptide analysis
• Sample processing in proteomics workflows

Pierce C18 Spin Tips offer excellent flow properties with a high-efficiency C18 sorbent for fast wetting, loading, washing and eluting. Sample is simply loaded in prepared tip and washed and eluted using multiple centrifugation steps. The procedure is simple and requires less than 5 minutes to process protein digests, strong cation exchange fractions and other protein and peptide samples for mass spectrometric (MS) analyses. The unique tip design also removes any particulates in the sample, eliminating clogging of the auto sampler or column upstream of the mass spectrometer.

Peptide Digest Assay Standard (1 mg/mL) (Thermo Scientific™)

Thermo Scientific™ Peptide Digest Assay Standard is a validated protein digest reference sample optimized for use in quantitative peptide assays to improve reproducibility and accuracy.

Robust peptide assay standard—the only commercially available peptide digest standard that has been developed for use in a quantitative peptide assay
Validated—validated for both the Pierce Quantitative Fluorometric Peptide Assay and the Pierce Quantitative Colorimetric Peptide Assay with comparable standard curves using both assays
Stable—liquid formulation is stored at 4⁰C, eliminating variability due to freeze-thaws
Easy-to-use—provided in a liquid format with a known peptide concentration

The Peptide Digest Assay Standard was designed as reference standard for use with the Pierce Quantitative Fluorometric and Colorimetric Peptide Assays to augment the reproducibility and accuracy of peptide quantitation in a microplate-based assay.

The reference protein has been digested with MS-grade trypsin to minimize missed cleavages. To help ensure consistent performance, digestion efficiency of the protein is monitored to help ensure lot-to-lot consistency, and quality is assessed by comparison to a reference standard. The Peptide Digest Assay Standard is provided in ready-to-use liquid format at 1 mg/mL, and is sufficient for at least ten standard curves (depending on dilution volumes).

Pierce™ Quantitative Colorimetric Peptide Assay (Thermo Scientific™)

The Thermo Scientific™ Pierce™ Quantitative Colorimetric Peptide Assay is an easy-to-use colorimetric microplate assay designed specifically to improve the sensitivity and reproducibility of quantitation of peptide mixtures.

Sensitive—accurately detect as little as 25 µg/mL of peptide mixture
Robust—assay performance rigorously tested using peptide digest mixtures
Robust peptide digest standard—kit includes a validated peptide digest standard for improved reproducibility of quantitation
Compatible—works with many reagents, including mass spectrometry sample preparation reagents and TMT™-labeled peptides
Convenient—easy mix-and read-format and stable colorimetric signal that may be read in as little as 15 minutes

The Pierce Quantitative Colorimetric Peptide Assay provides modified BCA reagents for the reduction of Cu+2 to Cu+1 and a proprietary chelator that have been optimized for the quantitation of peptide mixtures. In this reaction, the copper is first reduced by the amide backbone of peptides under alkaline conditions (Biuret reaction). The proprietary chelator then couples with the reduced copper to form a bright red complex with absorbance of 480 nm. The signal produced from this reaction is 3-4 fold more sensitive than the Micro-BCA Protein Assay for peptide analysis.

The kit also contains a high quality peptide digest reference standard for use in generating linear standard curves and to serve as a calibration control. This colorimetric peptide assay requires a small amount of sample (20 µL) and has a working peptide concentration range of 25–1000 μg/mL. The increased sensitivity, low sample assay volume, and included reference standard are essential for accurate and robust measurement of peptide digest samples, especially for mass spectrometry applications.

Applications:
• Recommended for peptide mixtures
• Not recommended for synthetic peptides because the assay is affected by peptide amino acid content
• Normalizing sample concentrations for quantitative downstream applications
• Normalizing sample loading volumes for LC, MS, and LC/MS applications
• Measuring recovery after peptide clean-up procedures

Pierce™ Quantitative Fluorometric Peptide Assay (Thermo Scientific™)

The Thermo Scientific™ Pierce™ Quantitative Fluorescent Peptide Assay is a sensitive, mix-and-read fluorescent microplate assay for the quantitative measurement of peptides and peptide mixtures.

Sensitive—accurately detect as little as 5.0 µg/mL of single peptides or peptide mixtures
Robust— assay performance rigorously tested using both peptides and peptide digest mixtures
Robust peptide digest standard—kit includes a validated peptide digest standard for improved reproducibility of quantitation
Compatible—works with many reagents, including those used in mass spectrometry sample preparation
Convenient—easy mix-and-read format and stable fluorescent signal that may be read in as little as 5 minutes up to several hours

The Pierce Quantitative Fluorescent Peptide Assay reagents include peptide assay buffer, fluorescent peptide labeling reagent, and a peptide digest assay standard for the quantitative measurement of peptide concentrations. In this assay, peptides are specifically labeled at the amino-terminus using an amine-reactive fluorescent reagent, and the fluorescently labeled peptides are detected using ex390/em475.

Because of the labeling mechanism of the fluorescent assay reagent, this assay is suitable for the quantitative measurement of synthetic peptides as well as peptide digest mixtures. A high quality peptide digest reference standard is included for use in generating linear standard curves and as a calibration control for improved accuracy and reproducibility. This sensitive assay requires only 10 µL of sample, produces a linear response with increasing peptide concentrations (5–1000 μg/mL), and results in a stable final fluorescence that can be detected in as little as five minutes.

Applications:
• Recommended for use with peptide mixtures and synthetic peptides with an unblocked amino-terminus (amino-terminus-labeled peptides, such as TMT-labeled peptides, are not compatible with this assay)
• Normalizing sample concentrations for quantitative downstream applications
• Normalizing sample loading volumes for LC, MS, and LC/MS applications
• Measuring recovery after peptide clean-up procedures.

Pierce™ Peptide Retention Time Calibration Mixture (Thermo Scientific™)

15 synthetic heavy peptides mixed in equimolar ratio to test and calibrate reverse-phase liquid chromatography (LC) and mass spectrometry (LC-MS) equipment.

Pierce™ TiO2 Phosphopeptide Enrichment Spin Tips (Thermo Scientific™)

Pierce TiO2 Phosphopeptide Enrichment Spin Tips are 200µL pipette tips containing TiO2 resin. This product is also sold as part of the Pierce TiO2 Phosphopeptide Enrichment and Clean-up Kit.

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Pierce™ TiO2 Phosphopeptide Enrichment and Clean-up Kit

Pierce™ MS-Compatible Magnetic IP Kit, protein A/G (Thermo Scientific™)

The Thermo Scientific™ Pierce™ MS-Compatible Magnetic IP Kit (Protein A/G) provides mass spectrometry-friendly reagents and an optimized protocol to enable highly effective and efficient immunoprecipitation and co-immunoprecipitation of target antigens upstream of LC-MS analysis.

MS-compatible—reagents directly compatible with in-solution peptide digestion, enriched samples contain minimal detergent residuals detected using LC-MS
Sensitive—procedure successfully enriches low abundance proteins (low ng range)
Low background—binding, wash, and elution buffers optimized to minimize enrichment of background proteins
Robust—procedure and reagents have been robustly tested with numerous targets to ensure consistent enrichment of low abundance proteins (ng range) with at least 2 peptides identified per protein

The Pierce MS-Compatible Magnetic IP Kit (Protein A/G) uses high-quality Pierce Protein A/G Magnetic Beads to provide wider flexibility of antibody capture than either Protein A or G alone. The optimized kit components have been formulated to be compatible with downstream LC-MS analysis. After the immunoprecipitation procedure, the target-enriched elution fraction is ready for in-solution tryptic digestion without the need for gel purification, detergent removal, or desalting.

This kit has been rigorously validated using numerous target antigens with varying expression levels, including targets previously undetected without enrichment or by western blotting. Additionally, the reagents and procedures have been validated using both manual and automated magnetic separation.

Applications:
• Enrichment of low abundance targets to improve MS sensitivity
• Identification of target proteins that are not detectable by western blotting
• Quantitation of target proteins by LC-MS analysis
• Screening of antibodies for IP-MS
• Enrichment of target proteins upstream of targeted analysis (SRM)

Pierce™ C18 Tips, 100 µL bed (Thermo Scientific™)

Thermo Scientific Pierce C18 Tips are ready-to-use pipette-tip columns of C18 resin that enable fast and efficient capture, concentration, desalting and elution of peptides for MALDI mass spectrometry and other methods.

Improve protein analysis results with Pierce C18 Tips by removing urea, salts and other contaminants before MS analysis. The tips are ideal for preparing samples for matrix-assisted laser desorption ionization (MALDI) or nanoelectrospray ionization techniques. The Pierce C18 Tips are available in convenient 10 and 100 µL tips with binding capacities of 8 and 80 µg, respectively.

Features of C18 Tips:

• Monolithic C18 fast-flow tips have low resistance and improved flow characteristics compared to other commercially available tips
• High binding capacity in a convenient tip format
• Two pipette tip formats for processing up to 8 or 80 µg of sample
• Eight packs and 96-tip trays are automation compatible

These C18 tips offer excellent flow properties with a high-efficiency monolithic C18 sorbent for fast wetting, loading, washing and eluting. The five-step procedure is simple and requires less than 5 minutes to process protein digests, strong cation exchange fractions and other protein and peptide samples for mass spectrometric (MS) analyses.

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Pierce™ C18 Tips, 10 µL bed

Pierce™ High pH Reversed-Phase Peptide Fractionation Kit (Thermo Scientific™)

Thermo Scientific™ Pierce™ High pH Reversed-Phase Peptide Fractionation Kit increases protein identification from LC/MS analysis through orthogonal peptide fractionation of complex peptide samples.

Easy-to-use—resin provided in single-use spin column format
Improved protein identifications—protein identifications increased by ≥50% when compared to unfractionated samples
Reproducible—elution profiles and fractional resolution vary by less than 20%
Optimized—robust procedure for maximal protein identification and peptide recovery while minimizing fractional overlap
Compatible—reagents validated with a variety of complex samples, including TMT™-labeled peptides

In order to enable deep proteome sequencing, it is often necessary to reduce the sample complexity by fractionation in an orthogonal dimension prior to LC/MS analysis. The Pierce High pH Reversed-Phase Peptide Fractionation Kit utilizes high pH reversed-phase chromatography to separate peptides by hydrophobicity and provides excellent orthogonality to low pH reversed-phase LC-MS gradients.

The kit has been designed to improve protein identification through the use of a proprietary reversed-phase resin in an easy-to-use spin column format with a highpH fractionation protocol. In contrast to strong cation exchange (SCX) fractionation, the high-pH reversed-phase fractions do not require an additional desalting step before LC/MS analysis.

The High pH Reversed-phase Peptide Fractionation Kit includes a high pH buffer (0.1% triethylamine) and twelve spin columns containing pH-resistant reversed-phase resin. Each reversed-phase fractionation spin column enables fractionation of 10–100 µg of peptide sample using a microcentrifuge.

Native, phosphorylated, Tandem Mass TagTM (TMTTM)-labeled, and other complex peptide mixture samples can be fractionated using this kit. Combining the search results generated by the individual fractions helps improve protein sequence coverage and increases the number of identified proteins relative to unfractionated samples.

Applications:
• Reducing sample complexity to identify targets of interest
• Conducting systems biology studies
• Reducing sample complexity to improve quantitation studies

Pierce™ MS-Compatible Magnetic IP Kit, streptavidin (Thermo Scientific™)

The Thermo Scientific™ Pierce™ MS-Compatible Magnetic IP Kit (Streptavidin) provides mass spectrometry-friendly reagents and an optimized protocol to enable highly effective and efficient immunoprecipitation and co-immunoprecipitation of target antigens upstream of LC-MS analysis.

MS-compatible—reagents directly compatible with in-solution peptide digestion, enriched samples contain minimal detergent residuals detected using LC-MS
Sensitive—procedure successfully enriches low abundance proteins (low ng range)
Low background—binding, wash, and elution buffers optimized to minimize enrichment of background proteins
Reduced antibody contamination—antibody biotinylation reduces contamination in the eluate when compared to other IP methods
Robust—procedure and reagents have been robustly tested with numerous targets to ensure consistent enrichment of low abundant proteins (ng range) with at least 2 peptides identified per protein

The Pierce MS-Compatible Magnetic IP Kit (Streptavidin) uses high-quality Pierce Streptavidin Magnetic Beads and optimized buffers that are compatible with downstream LC-MS sample preparation and analysis. After the immunoprecipitation procedure, the target-enriched elution fraction is directly compatible with in-solution tryptic digestion, eliminating the need for gel purification, detergent removal, or desalting.

This kit has been rigorously validated using numerous target antigens with varying expression levels, including targets previously undetected without enrichment or by western blotting. To support IP and co-IP applications using streptavidin supports, the Pierce Antibody Biotinylation Kit for IP has been specifically designed to enable biotin labeling of antibodies. Additionally, the reagents and procedures have been validated using both manual and automated magnetic separation.

Applications:
• Enrichment of low abundance targets to improve MS sensitivity
• Identification of target proteins that are not detectable by western blotting
• Quantitation of target proteins by LC-MS analysis
• Enrichment of target proteins from plasma samples for MS analysis

Pierce™ C18 Tips, 10 µL bed (Thermo Scientific™)

Thermo Scientific Pierce C18 Tips are ready-to-use pipette-tip columns of C18 resin that enable fast and efficient capture, concentration, desalting and elution of peptides for MALDI mass spectrometry and other methods.

Improve protein analysis results with Pierce C18 Tips by removing urea, salts and other contaminants before MS analysis. The tips are ideal for preparing samples for matrix-assisted laser desorption ionization (MALDI) or nanoelectrospray ionization techniques. The Pierce C18 Tips are available in convenient 10 and 100 µL tips with binding capacities of 8 and 80 µg, respectively.

Features of C18 Tips:

• Monolithic C18 fast-flow tips have low resistance and improved flow characteristics compared to other commercially available tips
• High binding capacity in a convenient tip format
• Two pipette tip formats for processing up to 8 or 80 µg of sample
• Eight packs and 96-tip trays are automation compatible

These C18 tips offer excellent flow properties with a high-efficiency monolithic C18 sorbent for fast wetting, loading, washing and eluting. The five-step procedure is simple and requires less than 5 minutes to process protein digests, strong cation exchange fractions and other protein and peptide samples for mass spectrometric (MS) analyses.

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High-Select™ TiO2 Phosphopeptide Enrichment Kit (Thermo Scientific™)

The Thermo Scientific™ High-Select™ TiO2 Phosphopeptide Enrichment Kit enables efficient enrichment of phosphorylated peptides from complex and fractionated protein digests for analysis by mass spectrometry (MS). (This product replaces Cat. No. 88301.)

Features of the TiO2 Phosphopeptide Enrichment Kit:
• Convenient—pre-formulated buffers for ease of use and spin-tip format to enable parallel processing of multiple samples
• High binding capacity—each column enriches up to 150 μg of phosphopeptides
• High specificity—recovers phosphopeptides with > 90% selectivity
• Complementary—TiO2 enriches a unique set of phosphopeptides that complements the High-Select™ Fe-NTA IMAC Phosphopeptide Enrichment Kit

The complete kit includes 24 titanium dioxide (TiO2) spin tips with optimized buffers to facilitate preparation of enriched phosphopeptides. Each column can enrich phosphopeptides from 0.5 to 3 mg of total protein digest as starting sample. The optimized buffers and simplified procedure eliminate the need for pyrrolidine in the elution buffer and subsequent clean-up using graphite spin columns. Use of this kit results in enrichment of up to 150 µg of phosphopeptides with greater than 90% selectivity.

Mass spectrometry is a key tool for identifying sites of protein phosphorylation and quantifying phosphorylation changes; however, MS analysis of protein phosphorylation is challenging due to the low stoichiometry, high hydrophilicity, poor ionization, and incomplete fragmentation of phosphopeptides. Because of the low relative abundance of phosphorylation modifications in complex protein samples, enrichment is essential for successful MS analysis of phosphopeptides. The improved High-Select TiO2 Phosphopeptide Enrichment Kit complements our lysis, reduction, alkylation, and digestion reagents, along with our C18, graphite spin, and high-pH reversed-phase peptide fractionation columns to provide a complete workflow for phosphopeptide enrichment.

The new procedure and optimized buffers have been shown to maintain the excellent selectivity and yield of the previous kit (Figure 1), while removing hazardous reagents and clean-up. The High-Select TiO2 Phosphopeptide Enrichment Kit is complementary to the High-Select Fe-NTA Phosphopeptide Enrichment Kit, with both kits enriching similar numbers of phosphopeptides but different subpopulations of phosphorylated peptides (Figure 2).

For high resolution analysis of the enriched phosphopeptides, the recommended LC column for Nanospray Flex source is Acclaim PepMap 100 C18 LC Column (Cat. No. 164942 or 164939). For EASY-Spray source, the recommended LC column is EASY-Spray C18 LC Column (Cat. No. ES802 or ES803).