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Identify microglia by using these antibodies, which are designed to detect protein biomarkers specifically expressed within these cells.
Streamline your neurodegeneration therapeutic development with CST recombinant monoclonal antibodies, ELISA and cellular assay kits, custom products, and services.
Learn about host-based and dye-conjugated methods for IF multiplexing, as well as a sequential labeling strategy that works with indirect detection.
Non-neuronal cells like microglia and astrocytes play critical roles in maintaining proper neuronal function, development, and disease.
Through in-house testing, we have found that the magnetic beads from our PTMScan HS kits aggregate and stick to the walls of some specific brands of tubes. Moreover, they cannot be washed off or spun off the sides of the tubes. This is an issue with specific brands of tubes themselves and not an inherent issue with the PTMScan bead product. Therefore, the following brands of polypropylene tubes are NOT recommended with magnetic bead PTMscan HS kits: 1. CellTreat #229441, polypropylene 1.5mL tube, non-sterile, non-pyrogenic2. Denville Scientific (Harvard Bioscience) #C3171, 1.5mL Posi-Lock tubes, polypropyleneThe following brand of microcentrifuge tubes is compatible with magnetic bead PTMscan HS kits:1. USA scientific tubes #1615-5500
Control peptides can be used to ensure reproducibility in proteomics experiments conducted with PTMScan.
The PTMScan® HS Ubiquitin/SUMO Remnant Motif (K-ε-GG) Kit #59322 uses the same antibody as in the traditional PTMScan Ubiquitin Remnant Motif (K-ε-GG) Kit #5562 and PTMScan Pilot Ubiquitin Remnant Motif (K-ε-GG) Kit #14482. This means it has the same affinity for the K-e-GG remnant after trypsin cleavage.However, we have changed the antibody – bead linkage to be near-covalent in strength so the antibody does not contaminate the PTM peptides during the elution step. This prevents issues with the liquid chromatography column clogging and increases the yield of PTM peptides identified. The specificity is also improved with changes to the bind and wash buffers that help reduce non-specific peptide binding.In addition, the new HS kit beads are magnetic, rather than agarose, which simplifies the workflow by eliminating multiple centrifugation steps during the bead washing and elution steps.
The PTMScan HS Magnetic Immunoaffinity Beads should not be frozen. We recommend storing this product at 4C. However, we have performed functional testing by IAP and subsequent mass spec analysis after 1X round of freeze thaws and found that the beads performed as expected. If the beads are accidentally frozen and thawed once, they should be fine to use, but we first recommend submitting a technical support inquiry to discuss this with a CST product scientist.
Antibodies and beads in the PTMScan® HS Ubiquitin/SUMO Remnant Motif (K-ε-GG) Kit #59322 are conjugated such that the antibody will not elute from the bead during peptide elution. The exact method of conjugation is proprietary.
PTMScan Motif Antibody Kits from Cell Signaling Technology
The incubation time for PTM peptide enrichment has been empirically determined with an optimum time of ~2 hours. Incubation times greater than 2 hours show no increase in PTM-specific enrichment, and shorter times may produce unwanted variability in the qualitative results.
The background binding of non-PTM peptides is typically 50% or more of the total peptides bound to the antibody beads in our agarose PTMScan® kits. This has been improved to less than 50% background or non-target PTM peptides for the magnetic PTMScan® HS kits. Therefore, it is hard to say from sample to sample exactly how much binding is from the antibody and how much is from the background. It is important to use a high-capacity stage tip that can handle 10ug of peptide at the end of the protocol after the enrichment step. In addition, we usually run 33% of each enrichment 2X for technical replicates for each sample.
Streamline Targeted Protein Degradation Drug Discovery with specific, application-validated antibodies and industry-leading proteomic services.
Discover the insulin signaling pathway and its role in glucose metabolism & diabetes. Learn more about insulin action here.
We recommend using 10 mg of protein per immunoaffinity purification (IAP) sample for our standard PTMScan kits. For the PTMScan HS product line, 1 mg of starting material is optimal. The minimum sample amount is dependent upon the cell line/tissue and the endogenous levels of post-translational modifications (PTMs). If less protein is used, fewer identifications could be expected.
We have observed >1E6 peak area for each control peptide when used at the supplied concentration followed by PTMScan and PTMScan HS enrichment, as detected by our LCMS instruments. However, the limit of detection (LOD) depends on many factors such as the sensitivity of the instrument and the LC gradient that is unique to each user. Users may adjust the concentration of the supplied peptides as needed to find an appropriate signal strength that has a sufficiently high signal-to-noise ratio for reliable detection.
The “classic” PTMScan® kits that use agarose beads and the PTMScan HS kits that use magnetic beads are both designed for a single PTM peptide enrichment. The nature of the acidic elution step, which leads to the release of bound PTM peptides, denatures the antibodies. This denaturing step alters the conformation of the antibodies. Therefore, employing these antibodies for subsequent enrichments may produce inconsistent results in PTM peptide enrichment.
What if an antibody could identify a PTM, regardless of the flanking amino acid sequence? This innovative idea is the basis of CST's PTMScan product line.
Lymphoma is a cancer of the lymphatic system, a key part of the body’s immune system. Treatment options & patient prognoses differ based on lymphoma type.
CUT&Tag delivers quality NGS data even if the signal from the Bioanalyzer or TapeStation System is low and the DNA library yield is low.
The Histone Modification Table provides a referenced list of many known histone modifications, associated modifying enzymes, and proposed functions.
The Ubiquitin Ligase Table provides a comprehensive list of E3 ubiquitin ligases along with their substrates and corresponding PubMed reference(s).
Learn how CUT&Tag cuts down your chromatin profiling experiment time. You’ll get the same great data with a fraction of the DNA library prep time and improved signal-to-noise.
The PI3K / Akt Substrates Table provides a comprehensive list of demonstrated downstream targets of Akt phosphorylation.
A list of antibodies directed against chaperones and other proteins involved in Protein Folding offered by Cell Signaling Technology.
A scientific resource for the GYF protein domain containing information on structure, function, and domain binding to proline-rich motifs.
This video describes the simplified PTMScan® HS workflow for LC-MS analysis of ubiquitination and SUMOylation that improves readout sensitivity, reduces sample prep time, and eliminates lyophilization, 9M urea, and antibody elution steps.
Alphabetical listing of protein, pathway, and antibody acronyms, curated by Cell Signaling Technology.
Clustered regularly interspaced short palindromic repeats, or CRISPRs, are repetitive sequences (direct repeats) native to the prokaryote genome.
