Technical Support Articles
Results (695)
What is the best way to choose an antibody for my CUT&RUN assay if I cannot find a CUT&RUN validated antibody for my target protein?
When selecting an antibody for use in CUT&RUN, we recommend starting with a ChIP or ChIP-seq-validated antibody; however, we have found that not all ChIP-validated antibodies work in the CUT&am...
What is the most suitable control DNA sample for my CUT&RUN NG-seq experiment?
When performing CUT&RUN with downstream NG-seq analysis, one can use a normal IgG antibody enriched sample as the negative control. However, we have seen and heard from other scientists that norma...
How did you determine that a NG-seq sequencing depth of 3 to 5 million sequencing reads was sufficient for CUT&RUN analysis?
This decision was based on our analysis of hundreds of CUT&RUN samples. We found that a sequencing depth lower than 3 million generated too few peaks and that the number of peaks increased with de...
How much CUT&RUN DNA is required for NG-seq analysis?
The amount of CUT&RUN DNA required for NG-seq analysis depends on the sensitivity of the DNA library prep kit. The typical yield for CUT&RUN DNA is 0.6 to 6 ng per reaction. Our We recommend using our SimpleChIP® ChIP-seq DNA Library Prep Kit for Illumina® #567... We have had customers perform this assay using frozen cell pellets, prior to adding any fractionation buffer, and the subsequent fractionation worked fine. Once you start fractionating, we recommen... We have several monoclonal antibodies for detecting total Akt protein that are validated for IHC-P in human samples. These are as follows:
What DNA library prep kit do you recommend using for CUT&RUN?
Can the Cell Fractionation Kit #9038 be used for frozen cell pellets?
Which total Akt antibody do you suggest for paraffin-embedded immunohistochemistry (IHC-P) with human samples?
What are the specifications for the 96-well plate in the CST ELISA kits?
For the 96-well colorimetric PathScan® and FastScan® ELISA plates, the specification information can be found in the We have used LysC in conjunction with trypsin for TMT-labeling-based studies to reduce miscleavages. For TMT-based workflows, we recommend a higher LysC:substrate ratio of 1:100 compared to the 1:250 ... Our CUT&RUN Assay Kit #86652 is compatible with downstream qPCR analysis. The pr... No, we don't recommend performing size selection during the library purification for CUT&Tag. Based on our experience and feedback from customers, size selection can significantly decrease bot... We recommend using DNA Purification Buffers and Spin Columns (ChIP, CUT&RUN, CUT&Tag) #14209 when purifying your CUT&Tag DNA. Although phenol-chloroform extraction followed by ethanol prec... For CUT&Tag assays using adherent cell lines, the first step is to detach the cells from the dish. We recommend using trypsin to detach the cells. Alternatively, Accutase Cell Dissociation Reagent... We have several monoclonal antibodies for detecting total Akt protein that are validated for IHC-P with mouse samples. These are as follows:
Can I use PTMScan® LysC Protease #84748 for tandem mass tag (TMT) labeling?
Can CUT&RUN enriched DNA be analyzed by qPCR, or do I have to do NGseq to analyze my data?
Do you recommend performing size selection of the DNA during the library preparation for CUT&Tag experiments?
Which method is better for purifying CUT&Tag DNA, DNA affinity columns or a phenol-chloroform DNA extraction using ethanol precipitation?
What advice do you have for collecting samples of adherent cell lines versus suspension cell lines for CUT&Tag experiments?
Which total Akt antibody do you suggest for paraffin-embedded immunohistochemistry (IHC-P) with mouse samples?
What is the fewest number of cells I can use in a reaction with the CST CUT&Tag kit?
We have shown that our CUT&Tag Assay Kit #77552 works with as few as 5,000-10,000 cells for histone modifications and 20,000 cells for transcription factors and cofactors.
Do I need to perform Fc blocking when using rabbit or mouse antibodies in flow cytometry?
Human Fc-gamma Receptor 1a (FCGR1A) exhibits strong cross-reactivity with rabbit IgG, and mouse IgG2a, IgG2c, and IgG3. Of course, mouse Fc-gamma Receptors will react with mouse antibodies. In flow cy...
How are TrkA, TrkB, and TrkC activated?
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...
Which S6 ribosomal protein antibody do you suggest for paraffin-embedded immunohistochemistry (IHC-P) with human samples?
We have several antibodies for detecting the phosphorylation of ribosomal protein S6 (rpS6) at Ser235/236 that are validated for IHC-P with human samples. These are S6 Ribosomal Protein (5G10) Rabbit ...
Which p44/42 MAPK (Erk1/2) antibody do you suggest for paraffin-embedded immunohistochemistry (IHC-P) with human samples?
We have several monoclonal antibodies for studying p44 and p42 mitogen-activated protein kinases (MAPKs), also referred to as Erk2 and Erk1, that are validated for IHC-P with human samples. These are ...
Why does the observed size versus the theoretical size of TET2 differ by western blot?
TET2 is post-translationally modified (https://www.phosphosite.org/proteinAction.action?id=22594&showAllSites=true). Although the theoretical molecular weight is 223.8 kDa these modifications incr...
What is the extinction coefficient of my rabbit antibody?
The molar extinction coefficient of rabbit IgG at 280 nm is approximately 210,000 M-1cm-1.
Why is the PTMScan® LysC Protease #84748 digestion performed at room temperature and not at 37°C?
The digestion is performed at room temperature to minimize carbamylation (also known as carbamoylation) of the sample peptides in the urea buffer, which can interfere with downstream PTMScan enrichmen...
Can CUT&Tag-enriched DNA be analyzed by qPCR, or do I have to do NGS to analyze my data?
If qPCR analysis is desired, we recommend performing CUT&RUN, as CUT&Tag DNA is not compatible with qPCR. The sample incubation step at 58℃ required for tagmented DNA solubilization in the ...
Why does the material in my new vial of Echinomycin #51434 look different?
Echinomycin #51434 is lyophilized into the vial, and any difference in the appearance of the lyophilized echinomycin is due to a dry coating of the compound in the vial. When you add dimethylsulfoxide...
Do you have mouse-reactive antibodies available for SignalStar™ Multiplex IHC?
Yes, please select “Mouse” in the first step of the SignalStar Multiplex IHC Panel Builder ...
My items say to store at -20°c but they were shipped in an envelope. Is it ok to use?
Yes, it is safe to use these items. Since 2003, CST has been shipping antibodies at room temperature when possible to reduce waste from packing materials. All CST antibodies are stability tested by ex...
When comparing my SignalStar™ staining to the chromogenic staining on a serial section, I see more positive cells. How do I know if this excess staining is correct?
During the course of optimization, we’ve found that fluorescent staining may show higher %-positivity than chromogenic staining. To ensure any excess staining is specific, confirm that the correct sub...
What is an appropriate positive control to include in this SignalStar™ assay? Are multiple controls necessary?
Any tissue shown to be positive for each marker via chromogenic IHC can serve as a positive control tissue for a SignalStar™ assay. Each target will therefore require a positive control, which may som...
I don’t see my target of interest in your menu of available antibodies for SignalStar™ Multiplex IHC. Can I still use it in my panel in some way?
SignalStar Multiplex IHC kits and reagents haven’t yet been validated for use with antibodies outside of our menu. We’re in the process of developing cust...
How long after the completion of staining can I wait to image my slides when performing a SignalStar™ Multiplex IHC assay?
For Imaging Round 1, the staining should show robust signal when imaged up to 8 hours post completion of staining. For Imaging Round 2, imaging should be performed as close to the completion of staini...