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When using antibodies for CUT&RUN against histone modifications, we typically see fragment sizes as small as 150 bp (size of a mono-nucleosome) and quite often see a nucleosomal ladder of DNA fragment sizes (i.e. 150, 300, 450, 600 bp). When using antibodies against transcription factors, we see a range of DNA fragment sizes, with the majority being greater than 35 bp.
Learn here about the HIF-1 signaling pathway and its impact on oxygen homeostasis and cellular adaptation. Click here.
Information and case study data for the PTMScan Direct Tyrosine Kinases Service from Cell Signaling Technology.
Profile protein abundance in cells and tissues using multiplexed sample labeling with Tandem Mass Tags and LC-MS/MS.
The family of Trk receptor tyrosine kinases consists of TrkA, TrkB, and TrkC. While the sequence of these family members is highly conserved, they are activated by different neurotrophins: TrkA by NGF, TrkB by BDNF or NT4, and TrkC by NT3.
The antigenic peptides used to produce our IRAK4 Antibody #4363 and our IRAK-M Antibody #4369 have been carefully selected to avoid cross-reactivity with the other IRAK family members. Our IRAK-M Antibody #4369 can detect a faint band at approximately 55 kDa. However, although the IRAKs are closely related in sequence, this band is not IRAK4.
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.
The PTMScan® technology is compatible with samples derived from all kingdoms of life as well as samples containing mixed species like xenografts (mouse and human), microbiome samples (bacterial and host species), and cells or tissue infected with pathogens. The only requirements are that the genome of the organisms be sequenced and the predicted open reading frames for the expressed proteins from the species under investigation be in the FASTA file format. This allows the search engine(s) performing peptide identification to match the experimentally generated MS2 chromatograms to those predicted by in silico chromatograms.
We suggest a sequencing depth of 3-5 million reads per sample. Because of the very low background signal generated in CUT&RUN, this sequencing depth is usually sufficient for histone modifications and transcription factors. We have found that less than 3 million does not work and greater than 15 million greatly increases the number of duplicate reads.
We work primarily with cell culture samples and do not typically test our ELISA kits with blood, serum, or other biological fluid samples. Depending on your kit of interest, it is possible human blood will be compatible. However, we cannot guarantee the performance.
The pH of our SignalStain® Antibody Diluent #8112 has a range between 7.38-7.45.
The additional amino acids found in SARS-CoV-2 Spike RBD (multimeric) (319-591) Recombinant Protein (8xHis-Tag) #17862 should not affect the binding of RBD to ACE2 because the RBD region ends at amino acid 527.
The Senescence β-Galactosidase Activity Assay Kit (Fluorescence, Plate-Based) #23833 is a relative quantitation assay, as it does not have a standard. Typically, researchers will first normalize their samples according to protein amount or cell number and then determine relative target abundance by comparing signal intensities between samples. We suggest customers run a control, such as a (-) treatment sample, in order to normalize their sample (+) treatment.
Although we have performed extensive testing, we have not found an appropriate model to detect phospho-DRP1 (Ser637) in human samples. DRP1, when phosphorylated at serine 637, is very difficult to detect, as it is rapidly degraded under most conditions. The region of the DRP1 protein surrounding serine 637 is conserved between rat and human, therefore we would predict our phospho-DRP1 (Ser637) antibodies to detect the human protein. However, since we have not been able to obtain signal in human samples, we cannot validate or guarantee these antibodies for use with human samples.
The higher band obtained with the TFE3 Antibody #14779 at approximately 80 kDa represents the sumoylated form of the TFE3 protein, which migrates higher than the unmodified form [see Miller, A.J. et al. (2005) J Biol Chem 280, 146-55 (PMID: 15507434; https://www.ncbi.nlm.nih.gov/pubmed/15507434)].
Both the polyclonal SS18-SSX Antibody #70929 and the SS18-SSX (E9X9V) XP® Rabbit mAb #72364 were produced by immunizing animals with a synthetic peptide corresponding to residues surrounding the fusion site of human SS18-SSX protein. There are two SS18 isoforms that can be fused to SSX, hence the presence of two bands.The SS18 (D6I4Z) Rabbit mAb #21792 also recognizes two distinct bands, corresponding to the two SS18 wild-type isoforms. The antigen used to produce the SS18 (D6I4Z) Rabbit mAb #21792 is downstream of the SS18 sequence (1-379) present in the fusion proteins and therefore #21792 does not detect the SS18-SSX fusions. Additionally, there are different SS18 fusions with SSX 1, 2, and 4; however, they are all the same size and based on sequence homology, both #70929 and #72364 should recognize all of them.We currently have two antibodies that recognize SSX proteins, SSX (E5A2C) Rabbit mAb (Carboxy-terminal Antigen) #23855 and SSX1/3 (E9P5M) Rabbit mAb (Amino-terminal Anti…
ChIP-seq is a powerful technique that combines ChIP and next-generation sequencing (NGS) to study DNA-protein interactions across the entire genome, but it is only as good as the quality of the antibody used in the ChIP experiment.
There is 50 µl of each antibody component that is listed in the StemLight Pluripotency Antibody Kit #9656. The kit components are pre-optimized for parallel use in immunofluorescent analysis. There are enough reagents for 100 assays based on a working volume of 100 µl. Another way to think of it is that if you need 100 µl of diluted antibody solution per sample/test, you will be using 0.5 µl of antibody (diluted into 100 µl) when used at the recommended 1:200 dilution.
When using overexpressed SOAT1 lysate or a SOAT1 enriched sample (i.e. immunoprecipitation) we observe multiple high molecular weight bands. Specifically for immunoprecipitation experiments, we speculate these bands are due to the experiment being performed in Cell Lysis Buffer (10X) #9803, which contains no SDS or DTT. Therefore, we believe the multiple high molecular weight bands are oligomers of SOAT1 (since they are 90, 135, 180 kDa vs the 45kDa monomer). These oligomers are formed under IP lysate conditions and cannot be broken up by SDS/DTT treatment after formation. In addition, when using normal endogenous cell lines which are lysed in SDS/DTT buffer we do not observe these high molecular weight bands.