Technical Support Articles

Why should I use the Protein A (HRP Conjugate) #12291 in place of the secondary antibody in my western blot for IP experiments?

Protein A predominantly binds intact IgG and does not bind denatured IgG well. It can be used in IP experiments to help avoid the potential masking of signal by denatured IgG heavy...

Will your GFP (D5.1) Rabbit mAb #2956 detect EGFP, eBFP2, and/or mCerulean3?

Our GFP (D5.1) Rabbit mAb #2956 is expected to detect EGFP, eBFP2, and mCerulean3.

Can I use human PBMCs with Senescence β-Galactosidase Staining Kit #9860?

CST scientists have not tested Senescence β-Galactosidase Staining Kit #9860 using human peripheral blood mononuclear cells (PBMCs). Therefore, we cannot provide a suggested...

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 containe...

Do your Phospho-Acetyl-CoA Carboxylase (Ser79) (D7D11) Rabbit mAbs detect both the alpha and beta isoforms?

Our Phospho-Acetyl-CoA Carboxylase (Ser79) (D7D11) Rabbit mAbs #11818 and #42801 detect both phospho-ACC-alpha and phospho-ACC-beta. These antibodies recognize ACC-alpha only when ...

Does your Anti-mouse IgG, HRP-linked Antibody #7076 cross react with mouse IgM?

Our Anti-mouse IgG, HRP-linked Antibody #7076 shows approximately 25% reactivity to mouse IgM.

What is your protocol for treating western blot membranes with CIP?

CIP-treated membranes are prepared with the following protocol:

REAGENTS:

Quick CIP - #M0525 (New England Biolabs)
rCutSmart™ Buffer - #B6004 (New England Biolabs)...

Is it possible to perform CUT&Tag on fixed frozen cells?

Yes. Cell fixation is recommended when the cells are very fragile (easily lysed during experiments), when the cellular signaling pathway is sensitive to the Concanavalin A beads...

Can you disclose the recipe for SignalStain® Antibody Diluent #8112?

While the formulation of our SignalStain® Antibody Diluent #8112 is considered proprietary, we can disclose that this product is PBS based and contains Tween-20 as well as BSA.

What DNA library prep kit do you recommend using for the CUT&Tag assay?

Which Cadherins does the Pan-Cadherin Antibody #4068 detect?

The Pan-Cadherin Antibody #4068 can detect R-, N-, E-, and VE-Cadherin.  It does not detect P- or K-Cadherin.  Reactivity to other Cadherins has not been tested.

What is the difference between phospho-Smad2 (Ser465/467) and phospho-Smad2 (Ser245/250/255)?

Phosphorylation of serines 465 and 467 indicates Smad2 activation and nuclear localization; whereas, phosphorylation of serines 245, 250, and 255 is inhibitory and leads to retenti...

What if my yield of CUT&Tag DNA is too low to see on an Agilent Bioanalyzer or TapeStation system?

Due to the nature of the Cell Fractionation Kit #9038 and the localization of each of the proteins in the Loading Control Sampler Kit #5142, the proteins will not be equally expres...

Why do you include BSA in the storage buffer for most of your off-the-shelf antibodies?

Antibodies, and proteins in general, are less susceptible to degradation when stored at higher concentrations (i.e., greater than or equal to 1.0 mg/ml). Therefore, because our off...

Do you observe any differences in CUT&Tag assay performance among different cell lines?

We have performed CUT&Tag on dozens of cultured adherent and suspension cell lines, both human and mouse. So far, we have not noticed any significant difference in performance ...

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 ...

Which phospho-S6 ribosomal protein (Ser235/236) 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 Phospho-S6 Ribo...

Can I use the Cell Fractionation Kit #9038 with tissue samples?

Since the Cell Fractionation Kit #9038 is primarily meant to be used for cultured cell lines, the buffers have not been optimized for use with tissue samples. Therefore, the fracti...

How do I validate the PTM sites identified in PTMScan® experiments?

The validation of PTM sites may be accomplished in several ways. Often, the site of interest may be seen in the context of more than one peptide due to alternative forms caused by ...