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CST promotes alternative transportation through a variety of programs.
For PTMScan® enrichments, we recommend a liquid chromatography (LC) gradient length of 90 minutes and replicate injections for each enrichment. Therefore, four PTMScan enrichments will have a total of eight liquid chromatography-mass spectrometry (LC–MS)/MS runs. For the immobilized metal affinity chromatography (IMAC) based phosphopeptide enrichments, the sample complexity requires a longer gradient length of 120 minutes, also performed with replicate injections for each sample.
We do not perform enzymatic activity assays internally using the Cell Lysis Buffer (10X) #9803, so we are unable to guarantee that the enzymatic activity will be conserved. We routinely use this buffer in immunoassays such as western immunoblot, IP, and ELISA without issue.As a reference, the components of the Cell Lysis Buffer (10X) #9803 are provided below:20 mM Tris-HCl (pH 7.5), 150 mM NaCl, 1 mM Na2EDTA, 1 mM EGTA, 1% Triton, 2.5 mM sodium pyrophosphate, 1 mM beta-glycerophosphate, 1 mM Na3VO4, 1 µg/ml leupeptin
CST has an energy management plan to reduce our use of resources.
The CUT&Tag Assay Kit #77552 contains components that are packaged carefully and need to be stored at either room temperature (RT), 4℃, or -20℃. It's important to store each reagent at the appropriate temperature to ensure optimal functionality.However, if the entire kit is accidentally stored at -20℃, the Concanavalin A beads will lose some activity. If this happens, we recommend purchasing our standalone Concanavalin A Magnetic Beads and Activation Buffer #93569 for future experiments, especially when working with low abundance and/or weak DNA binding proteins. In addition, to preserve proper performance, thaw the DNA spin columns slowly (i.e., thaw at 4℃ first, then calibrate to room temperature) and make sure there is no condensation inside the column before use. If needed, DNA Purification Buffers and Spin Columns (ChIP, CUT&RUN, CUT&Tag) #14209 are also available for purchase as a standalone product. All of the other reagents in the kit are acceptable for use aft…
Sonication ChIP Kit Performance Comparison
Profile protein abundance in cells and tissues using multiplexed sample labeling with Tandem Mass Tags and LC-MS/MS.
We have not tested the reactivity of our myosin light chain 2 antibodies against myosin 12a/b (UniProt: P19105 and O14950). However, based on homology, there is a good chance for cross-reactivity. The length of MLC12a/b is 171 amino acids and 172 amino acids respectively and both are predicted to run at ~20 kDa.The peptide immunogens used to produce our Myosin Light Chain 2 antibodies have the following sequence homology with Myosin 12a and 12b:
Information and case study data for the PTMScan Direct Tyrosine Kinases Service from Cell Signaling Technology.
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.
We do not routinely test samples prepared from whole tissue. However, customers have successfully used our ELISA kits with their tissue extracts. If you are interested, we can provide a recommended protocol for tissue sample preparation. You will likely need to optimize the amount of tissue that you are loading because of the lower than normal concentration of the target of interest. These kits come with a positive control that can be assayed alongside the tissue sample.
As a company rooted in science, we are troubled by recent reports citing antibodies as one of the causes of scientific irreproducibility.
CST’s does not ship in styrofoam coolers and packaging materials are recyclable.
PEG (polyethylene glycol) has been added to our Anti-rabbit IgG (H+L) (DyLight™ 800 4X PEG Conjugate) #5151 and Anti-mouse IgG (H+L) (DyLight™ 800 4X PEG Conjugate) #5257 to increase their solubility and stability for various applications, including cellular and in vivo imaging. Internal Fluorescent Western and In-cell Western testing shows that the addition of PEG to these secondaries not only significantly increases signal but also decreases background fluorescence.
All of our siRNAs are single duplexes. We discontinued pools a number of years ago to avoid off-target effects.
We have confirmed in-house that our Phospho-p53 (Ser15) Antibody #9284 and Phospho-p53 (Ser15) (D4S1H) Rabbit mAb #12571 cross-react with some prestained protein markers. We are unsure as to why this is occurring. To help avoid this, we recommend using a biotinylated marker without the use of a prestained marker.
It is critical that our Phospho-p53 (Ser15) Antibody #9284 be diluted into 5% w/v non-fat dry milk, 1X TBS, 0.1% Tween® 20 and incubated at 4°C with gentle shaking, overnight. Diluting the antibody in BSA will result in high background on the blot.
Cell Lysis Buffer (10X) #9803 will interfere with the Bradford protein assay. This is because it contains detergent. To make the assay somewhat compatible, dilute your protein quantitation samples at least 10-fold with water or PBS to reduce the amount of detergent and consequentially lower the interference. This ten-fold dilution may still yield background absorbance. We recommend employing an alternative protein assay that is compatible with #9803, such as the BCA Protein Assay Kit #7780.
There are 3 isoforms of Estrogen Receptor α (ER66, ER46, and ER36). Our Estrogen Receptor α (D8H8) Rabbit mAb #8644 will recognize ER66 and ER46, but it will not recognize ER36.
It is possible to see a doublet at ~100 kDa for Grp94 due to post-translational modifications. Here is a link to our PhosphoSitePlus database detailing the possible post-translational modifications for Grp94: https://www.phosphosite.org/proteinAction.action?id=6928&showAllSites=true.
HPSE is synthesized as a 65 kDa inactive precursor that undergoes proteolytic processing, yielding 8 kDa and 50 kDa protein subunits that heterodimerize to form an active enzyme. The 65kDa proform and the 50 kDa subunit (amino acids 158-543) are detected by the HPSE (E3N3H) Rabbit mAb #99756, while the 8kDa subunit (amino acids 36-109) is not.
The predicted molecular weight of PD-L2 is 31 kDa (based on amino acid composition). However, in most models, PD-L2 migrates at an observed molecular weight of 45-60 kDa on Tris-Glycine gels. This discrepancy between the predicted and the observed molecular weights is a result of glycosylation [see Wang, H et al. (2017) Oncoimmunology 6(7), e1327494 (PMID: 28811964; https://www.ncbi.nlm.nih.gov/pubmed/28811964)].
We have several antibodies for β-actin that are validated for IHC-P in human samples. These are β-Actin (13E5) Rabbit mAb #4970 and β-Actin (8H10D10) Mouse mAb #3700. Both work well and there is no preference for this application.