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The Trusted NLRP3 Antibody for Western Blotting

AdipoGen Life Sciences' anti-NLRP3/NALP3, mAb (Cryo-2) is the Standard antibody to detect NLRP3 by Western blotting. With over 1000 citations and used by "the" experts in the inflammasome research field, this antibody is the first choice for immunoblotting of NLRP3.
It can also be used in ICC, IHC, IP and ChIP assays, as proven in various studies.
See our Website for NLRP3 Western blotting Tips and Protocols.

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Product Details
Synonyms NACHT-, LRR- and PYD-containing Protein 3; NLRP3; Cryopyrin; Cold Autoinflammatory Syndrome 1 Protein
Product Type Monoclonal Antibody
Properties
Clone Cryo-2
Isotype Mouse IgG2b
Source/Host Purified from concentrated hybridoma tissue culture supernatant.
Immunogen/Antigen Recombinant mouse NLRP3/NALP3 (pyrin domain/aa 1-93).
Application

Western Blot (see online protocol): (1μg/ml)
Immunocytochemistry: (see J. Virol. 85, 13019 (2011) and Nat. Immunol. 13, 255 (2012))
Immunoprecipitation: (in RIPA buffer, see Mol. Cell 49, 331 (2013))
Immunohistochemistry: (1:200, for a protocol see Nature 493, 67 (2012))
ChIP assay: 2µg/DNA of 10E6 cells (see Nat. Immunol. 16, 859 (2015))
Crossreactivity Human
Mouse
Specificity

Recognizes mouse and human NLRP3/NALP3.
Purity ≥95% (SDS-PAGE)
Purity Detail Protein G-affinity purified.
Concentration 1mg/ml
Formulation Liquid. In PBS containing 10% glycerol and 0.02% sodium azide.
Isotype Negative Control

Mouse IgG2b Isotype Control

Other Product Data

Tips for Western blotting: 

1. NLRP3 is found only at low levels and the tissue or cells should express enough NLRP3. Therefore, the priming and activation steps are crucial for a successful Western blot and are often the reason why NLRP3 can not be detected properly. Please make sure that you have primed and activated the NLRP3 inflammasome properly.

2. The main band detected with the antibody Cryo-2 is migrating at about 110kDa. For high molecular weight proteins >100kDa such as NLRP3, it is best to transfer overnight following the tips described below. If the NLRP3 is not well transferred, then a longer exposure time is required and other non-specific bands can appear. A semi-dry blot is usually not recommended for proteins higher than 100kDa. We recommend using liquid transfer (wet tank) at 20V overnight to be sure that all NLRP3 has been transferred. Important, if markers are well transferred to the membrane, it does not mean that NLRP3 is transferred well too (markers are selected to be easily transferred also the High Molecular Weight Markers).

For the NLRP3, it is best to apply the following tips:
- Please perform an immunoblotting experiment using a 6-7% SDS-PAGE gel.
- Use 15-20% methanol in the transfer buffer (except for PVDF).
- Do not let the gel sit in the transfer buffer for a prolonged period before transfer.
- Transfer for 16 hours (O/N) at 20 Volts.
- 0.45 µm PVDF or nitrocellulose may be effective.
- Make sure the transfer unit does not overheat during the electroblotting procedure.

3. We recommend using THP1 or BMDCs as a positive control and performing a 2nd antibody-alone control in parallel.
RRID AB_2490202
Shipping and Handling
Shipping BLUE ICE
Short Term Storage +4°C
Long Term Storage -20°C
Handling Advice After opening, prepare aliquots and store at -20°C.
Avoid freeze/thaw cycles.
Use/Stability Stable for at least 1 year after receipt when stored at -20°C.
Documents
Protocols Download PDF
MSDS Download PDF
Product Specification Sheet
Datasheet Download PDF

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