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PTMScan® Videos and Webinars

PTMScan videos and links to complementary technologies for phospho-peptide enrichment and Science Webinar on Proteomics and Cell Signaling.

Flow Cytometry (F) Protocol for the Detection of Transcription Factors

Flow Cytometry (F) Protocol for the Detection of Transcription Factors with recommended companion products.

CH Protein Domain

A scientific resource for the CH protein domain containing information on structure, function, and binding to phospho-serine and phospho-threonine motifs.

Resources

The recommended primary antibody dilution buffer for western blotting (i.e., BSA or nonfat dry milk) can be found either in a grey box on the bottom of the first page of the product datasheet or near the top of the product-specific protocol.  

The product-specific western immunoblotting protocol can be found under "Product Information" on each product's webpage: 

1) Enter the product number in the "Products" search box and hit enter.
2) Scroll down to "Product Information" on the product webpage.
3) Select "Western Blotting" in the drop-down menu under "Protocol".
4) Find BSA or nonfat dry milk listed directly under the "Western Blotting Protocol" heading.
 

What is glycolysis and what is its role in metabolism?

What is glycolysis? Glycolysis and oxidative respiration are the two metabolic pathways used to convert biochemical energy into ATP to fuel cell function.

Flow Cytometry Protocol for FoxP3 on Murine Splenocyte T Cells

Flow Cytometry Protocol for FoxP3 on Murine Splenocyte T Cells: easy to follow directions describing the step by step experimental procedure.

Organic Ideas for Composting at CST

We wanted our first innovation challenge to be based on something visible and measurable, so cup reduction and compost awareness were good baseline goals.

Stress Granules: What are they and what role do they play in neurodegenerative diseases?

Stress granules are a cellular defense mechanism against stress but disruption in disassembly can lead to toxic proteins and neurodegenerative diseases.

Webinar | Deciphering Cancer: Genomic Instability in Cancer

This webinar will examine how we may be able to develop effective new therapeutic options for cancer treatment by targeting proteins responsible for chromatin modifications.

Immunology: Which cells have a myeloid lineage and how are they identified?

Identifying and quantifying myeloid subtypes is essential for understanding why different cell populations are activated in response to certain pathogens.

Resources

Our Pan-Calcineurin A Antibody #2614 detects endogenous levels of the alpha isoform of Calcineurin A, however it may also recognize the beta and gamma isoforms. Regrettably, because it has been difficult to find suitable models, we have not specifically tested if this product will detect the beta and gamma isoforms. However, the antigenic peptide shares around 90% sequence identity with the beta isoform and around 80% sequence identity with the gamma isoform. Therefore, it is possible that there is cross-reactivity. This antibody does not cross-react with protein phosphatase 1 or 2A.

Resources

Unfortunately, we have not internally validated the use of acidic samples with our Cyclic AMP XP(R) Assay Kit #4339. We have had customers try this method, preparing their samples with 0.1M HCL and then neutralizing. The customers did not have success with this method.

If using acidic sample preparation, we recommend further diluting samples in 1X PBS after neutralization, prior to adding samples to the plate. Again, as we have not internally validated this, we cannot guarantee this kit will work with acidic samples. 

Resources

Yes, the flow through from a PTMScan experiment can be saved and stored at -80C for future enrichments. We have performed 5 different PTM enrichments sequentially from one sample. The limitation would be the choice of the initial protease used for the digest. For example, a UbiScan PTMScan requires peptides digested with trypsin and the flow through peptides may not be ideal to capture peptides that require an arginine (R) residue in the phospho-motif as in the AKT or AMPK substrate PTMScan kits.

WD40 Protein Domain

A scientific resource for the WD40 protein domain containing information on structure, function, and binding to both phospho-Ser/Thr and methylated lysine residues.

Resources

This product is purified from goat blood and is therefore 100% goat serum. If you are using it as a blocking buffer, you can dilute it as follows:

Blocking Buffer: 1X PBS + 5% normal serum + 0.3% Triton™ X-100
To prepare 10 mL, add 0.5 mL normal serum from the same species as the secondary antibody (e.g., Normal Goat Serum #5425 ) to 9.5 mL 1X PBS and mix well. While stirring, add 30 µL Triton™ X-100.

Resources

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 the Senescence β-Galactosidase Staining Kit #9860:

Reagent grade water
Potassium Hexacynoferrate (III) [CAS 13746-66-2]


 

UIM Protein Domain

A scientific resource for the UIM protein domain containing information on structure, function, and domain binding to ubiquitin.

Resources

X-Gal can be tricky to solubilize, therefore we recommend dissolving in DMF (#12767) as listed on the Senescence β-Galactosidase Staining Kit (#9860) datasheet. However, we have tested and verified that DMSO (#12611) can be used to successfully dissolve the X-Gal, and therefore we consider it to be a suitable replacement for DMF. 

Resources

The suggested protocol for treating cells with poly(dA:dT) can be found below: 

Poly(dA:dT) Treatment Protocol (5 μg/mL final concentration)

Reagents:

Procedure:

24 hours prior to transfection:
  • Plate cells in two 15 cm plates with 21 mL media with serum (no antibiotic)

On transfection day:

  1. Prepare two falcon tubes per plate to be transfected:
    • For mock: One tube labeled “FuGENE” and one tube labeled “Mock”
    • For poly(dA:dT): One tube labeled “FuGENE” and one tube labeled “Poly(dA:dT)”
  2. Add 4.5 mL of 1X serum-free medium to ea…

Reproducibility in Research Science - Join the Discussion.

The reproducibility of scientific studies has become a major issue, leading to a lack of trust in scientific results from the academic and pharmaceutical research communities.

Resources

With adherent cell lines, one first needs to detach the cells from the dish. We recommend using trypsin to detach the cells. Alternatively, you can use accutase, but the resulting cell pellet doesn't pack as well during centrifugation, so you tend to lose cells during the wash steps. Suspension cells are much easier. Just count the cells using a hemacytometer or auto cell counter and then collect the number of cells you need. We don't observe a significant difference in assay performance with adherent versus suspension cells.

ApoE4 and the Insulin Receptor — It's a Trap!

Richard discusses a recent paper that investigates the potential mechanistic link between the AD risk gene APOE4 and altered metabolism. This makes an interesting potential connection from diabetes to Alzheimer's.

VHL Protein Domain

A scientific resource for the VHL protein domain containing information on structure, function, and domain binding to hydroxylated proline residues.

Resources

The TGF-β Antibody #3711 detects recombinant TGF-β1, TGF-β2, and TGF-β3. The antibody also detects endogenous levels of the TGF-β1 precursor proteins. The different sizes we have listed represent the precursor TGF-β protein (45kDa), the mature dimer form (25kDa) and the monomer (12kDa).

We do not usually detect the mature TGF-β at 25 kDa because mature TGF-β is composed of two polypeptides that are disulfide-linked. Therefore under reducing conditions, these polypeptides are separated and observed as a single band at 12 kDa. However, we have shown that the TGF-β Antibody #3711 can detect the mature form of TGF-β by using recombinant protein.

TGF-β is rapidly exported from the cell and therefore may be difficult to detect in tissue or cell lysates. In human platelet samples, we observe two strong bands, with the larger band likely being the precursor (uncleaved) form at 45kDa and the smaller being the monomer at 12kDa.

UEV Protein Domain

A scientific resource for the UEV protein domain containing information on structure, function, and domain binding to ubiquitin.

BAR Protein Domain

A scientific resource for the BAR protein domain containing information on structure, function, and binding to phospholipids.

SH3 Protein Domain

A scientific resource for the SH3 protein domain containing information on structure, function, and domain binding to proline-rich motifs.

Writing a Grant Part 8: What Are My Chances?

We've hit on every aspect of grant writing in these blogs. You may now be wondering: what are my chances? Am I likely to get funded? We dig into the data.

BH1-4 Protein Domains

A scientific resource for the BH protein domains containing information on structure, function, and domain binding in apoptosis.

What Is Neurodegenerative Disease?

Neurodegeneration is marked by the progressive loss of neuronal function and structure, leading to cognitive disabilities such as dementia.