Technical Support Articles
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Does the amino acid sequence of SARS-CoV-2 Spike RBD (multimeric) (319-591) Recombinant Protein (8xHis-Tag) #17862 (319-591) being longer than the amino acid sequence of other RBD products (318-541) affect its binding activity with ACE2?
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 ...
Why does the signal for LC3-II increase with chloroquine, an inhibitor of autophagy?
The abundance of LC3-II is generally very low without treatment. Glucose starvation, amino acid starvation, serum starvation, and rapamycin treatment have been shown in the literature to induce increa...
What is your recommended procedure for antigen retrieval with a pressure cooker?
For antigen retrieval in a pressure cooker, bring slides submerged in buffer up to 125C, maintain for 30 seconds, then cool to 90C and maintain that for 10 sec. Remove the container from the device an...
Which beta-actin antibody do you suggest for paraffin-embedded immunohistochemistry (IHC-P) with human samples?
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 pr...
Which cleaved caspase-3 (Asp175) antibody do you suggest for immunofluorescent immunocytochemistry (IF-IC) with human samples?
We have several antibodies for detecting Cleaved Caspase-3 (Asp175) that are validated for IF-IC with human samples. These are:
- Cleaved Caspase-3 (Asp175) Antibody #9661
- Cleaved Caspas...
Does your CUT&RUN Assay Kit work with isolated nuclei?
Our CUT&RUN Assay Kit is optimized for use with whole cells and works with a large number of cell lines, both adherent and suspension. While it is not typically necessary to pre-isolate cell nucle...
What isoforms does the ZBP1 Antibody #60968 recognize?
ZBP1 (Z-DNA binding protein 1), also referred to as DAI (DNA-dependent activator of IFN-regulatory factors) and DLM-1, is a nucleotide-binding protein that plays a role in tumorigenesis and innate imm...
How should I resuspend and aliquot the PTMScan® LysC Protease #84748?
To resuspend the PTMScan® LysC Protease #84748, we recommend allowing the vial to equilibrate at room temperature for a few minutes after removing from the -20C freezer. Next, spin the vial at 1,0...
Can I use primers from the Illumina Nextera Library preparation system or design my own primers to amplify the DNA library generated from the CUT&Tag Assay Kit #77552??
Yes, you can use primers from the Illumina Nextera Library preparation system or self-designed primers to amplify the DNA library generated from the CUT&Tag DNA using the CUT&Tag Assay Kit #77...
Should I be concerned that my Concanavalin A beads "clump together" in some of my CUT&RUN experiments?
The Concanavalin A beads can clump together rather easily, especially when the cells become lysed. To avoid this, make sure your cells are healthy and be sure to treat them very gently during the cell...
Why is the concentration of my antibody so low?
At CST, our antibodies are formulated on the basis of their reactivity and performance for each approved application at a recommended dilution. For antibodies with high affinity, optimal performan...
Will your Anti-biotin, HRP-linked Antibody #7075 cross-react with endogenous biotin in cell lysates?
Our Anti-biotin, HRP-linked Antibody #7075 is intended for use with our Biotinylated Protein Ladder Detection Pack #7727. However, it is possible for this antibody to also detect endogenously bioti...
Is Fc blocking needed for F(ab') antibody fragments used in live cell flow cytometry assays?
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...
Does CUT&RUN show a bias toward euchromatin or heterochromatin?
We do not observe any bias toward euchromatin or heterochromatin in our assays. In CUT&RUN, the target-specific antibody recruits the pAG-MNase and directs digestion to chromatin that is directly ...
Which phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) antibody do you suggest for paraffin-embedded immunohistochemistry (IHC-P) with human samples?
We have several antibodies for detecting p44 and p42 MAPK when phosphorylated at Thr202 and Tyr204 of Erk1, or Thr185 and Tyr187 of Erk2, that are validated for IHC-P with human samples. These are Pho...
Which phospho-Akt (Ser473) antibody do you suggest for Immunofluorescence (Immunocytochemistry) with mouse samples?
We have several antibodies for detecting Akt phosphorylation at Ser473 that are validated for IF-IC with mouse tissues. These are:
- Phospho-Akt (Ser473) (D9E) XP® Rabbit mAb #4060
Why should I use your recommended protocol or reagents for my IHC experiment?
We have performed a number of comparisons over the years looking at protocol variables such as the retrieval solution, diluent, incubation, and detection methodology. We have seen that the choice of r...
What controls can I use to show my CUT&RUN experiment is working?
Whether performing CUT&RUN with downstream qPCR or NG-seq analysis, we always recommend using a CUT&RUN-validated positive control antibody like our We are currently unable to provide the Solution B component separately from the Senescence β-Galactosidase Staining Kit #9860. However, we can disclose the ingredients of Solution B that is used in th... Although LysC can digest proteins at high urea concentrations (~8M), we recommend diluted urea (~2M) to minimize the risk of carbamylation. Our CD45 (D9M8I) XP (#13917), CD45 (D4H7K) ( #72787), and CD45 (D3F8Q) (#70257) Rabbit mAbs recognize intracellular (cytoplasmic) epitopes in the human (UnitProt ID We recommend using a CUT&Tag-validated positive control antibody, like our Tri-Methyl-Histone H3 (Lys4) (C42D8) Rabbit mAb #9751, in your experiment to confirm that your CUT&Tag assay is worki... Our in-house threshold for a positive signal is an OD reading of 1.5. Each ELISA kit has a different lower and higher limit of signal detection, based on the sensitivity profile of each individual kit... For PTMScan experiments, the stage tip, or small-scale peptide desalting is a critical part of the experiment for a number of reasons. Detailed instructions for performing stage tip peptide desalting ... The suggested protocol for treating cells with poly(dA:dT) can be found below: Yes, our CUT&Tag Assay Kit works with flash frozen tissue samples or tissue samples directly frozen at -80℃. After thawing, fixation of the tissue sample is optional, as we haven't found that ... Occasionally, Cell Lysis Buffer (10X) #9803 stored at -20C may not appear frozen, which is not a reason for concern. This phenomenon is known as nucleation, or the initial formation of ice from a liqu... We have several antibodies for detecting ribosomal protein S6 (rpS6) that are validated for IF-IC with mouse samples. These are S6 Ribosomal Protein (5G10) Rabbit mAb #2217 and S6 Ribosomal Protein (5... The Senescence β-Galactosidase Staining Kit #9860 is intended to be a qualitative assay to determine if cells are senescent. We have not determined a means of quantification in-house. However, we usua...Can I purchase Solution B outside of the Senescence β-Galactosidase Staining Kit #9860?
Why is the PTMScan® LysC Protease #84748 digestion performed in a diluted urea formulation?
What CD45 isoforms are detected by the CD45 (D9M8I) XP (#13917), CD45 (D3F8Q) (#70257), and CD45 (D4H7K) ( #72787) Rabbit mAbs?
What positive and negative controls can I use to demonstrate that my CUT&Tag experiment is successful?
What are the lower and higher limits of detection for your ELISA kits?
Why should I include a stage tip step in my PTMScan® experiment?
Does CST have a protocol for treating cells with poly(dA:dT)?
Poly(dA:dT) Treatment Protocol (5 μg/mL final concentration)Is it possible to perform CUT&Tag on frozen tissue samples?
Why isn't my bottle of Cell Lysis Buffer (10X) #9803 frozen?
Which S6 ribosomal protein antibody do you suggest for immunofluorescence-immunocytochemistry (IF-IC) with mouse samples?
Can I quantify my samples using the Senescence β-Galactosidase Staining Kit #9860?