Cell Signaling Technology

Product Pathways - Protein Folding

HSP/Chaperone Antibody Sampler Kit #9965

Kit Includes Quantity Applications Reactivity MW (kDa) Isotype
HSP60 (D307) Antibody #4870 40 µl W IF-IC F H M R Mk Dm 60 Rabbit
HSP70 Antibody #4872 40 µl W IHC-P H M R Mk B 72, 73 Rabbit
HSF1 Antibody #4356 40 µl W IP IHC-P IF-IC F ChIP H M R Mk 82 Rabbit
BiP (C50B12) Rabbit mAb #3177 40 µl W IHC-P IHC-F H M 78 Rabbit IgG
HSP40 (C64B4) Rabbit mAb #4871 40 µl W IHC-P H M R Mk 40 Rabbit IgG
HSP90 (C45G5) Rabbit mAb #4877 40 µl W IHC-P IF-IC F H M R Mk (B) 90 Rabbit IgG
Calnexin (C5C9) Rabbit mAb #2679 40 µl W IHC-P IF-IC H Mk 90 Rabbit
PDI (C81H6) Rabbit mAb #3501 40 µl W IHC-P IF-IC H M R Mk 57 Rabbit
Anti-rabbit IgG, HRP-linked Antibody #7074 100 µl Goat

Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IHC-P=Immunohistochemistry (Paraffin)  IHC-F=Immunohistochemistry (Frozen)  IF-IC=Immunofluorescence (Immunocytochemistry)  F=Flow Cytometry  ChIP=Chromatin IP
Reactivity Key:  H=Human  M=Mouse  R=Rat  Mk=Monkey  Dm=D. melanogaster  B=Bovine
Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Specificity / Sensitivity

HSP40 (C64B4) Rabbit mAb detects endogenous levels of total HSP40 protein. HSP60 (D307) Antibody detects endogenous levels of total HSP60 protein. HSP70 Antibody detects endogenous levels of total HSP70 protein (HSP70-Hom, HSP70-1). HSP90 (C45G5) Rabbit mAb detects endogenous levels of total HSP90 protein. HSF1 Antibody detects endogenous levels of total HSF1 protein. Calnexin (C5C9) Rabbit mAb detects endogenous levels of total calnexin protein. PDI (C81H6) Rabbit mAb detects endogenous levels of total PDI protein. BiP (C50B12) Rabbit mAb detects endogenous levels of total BiP protein. Each of these antibodies recognizes only its specific target.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell types using Calnexin Antibody #2433.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell types using PDI Antibody #2446.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell types using BiP (C50B12) Rabbit mAb #3177.


Western Blotting

Western Blotting

Western blot analysis of extracts from various cell types using HSF1 Antibody #4356.

Western Blotting

Western Blotting

Western blot analysis of extracts from HeLa, C2C12 and C6 cells using HSP40 Antibody #4868.

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell types using HSP60 (D307) Antibody #4870.


Western Blotting

Western Blotting

Western blot analysis of extracts from various cell types using HSP70 Antibody #4872.

Western Blotting

Western Blotting

Western blot analysis of extracts from HeLa, NIH/3T3 and C6 cells using HSP90 (E289) Antibody #4875.

Description

The HSP/Chaperone Sampler Kit provides an economical means to investigate protein folding within the cell. The kit contains enough primary and secondary antibodies to perform four Western blot experiments with each antibody.

Source / Purification

Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to Asp307 of human HSP60, corresponding to human HSP70, and corresponding to residues at the carboxy-terminus of human HSF1 protein. Polyclonal antibodies are purified by protein A and peptide affinity chromatography. Rabbit monclonal antibodies are produced by immunizing rabbits with a synthetic peptide corresponding to residues surrounding Gly584 of human BiP, surrounding Asn300 of HSP90, corresponding to Glu223 of human HSP40/Hdj1, corresponding to the sequence of human calnexin, and corresponding to the sequence of human PDI.

Background

HSP70 and HSP90 are molecular chaperones expressed constitutively under normal conditions to maintain protein homeostasis and are induced upon environmental stress (1). HSP70 and HSP90 interact with unfolded proteins to prevent irreversible aggregation and catalyze the refolding of their substrates in an ATP-dependent manner (1). HSP40 family proteins bind unfolded proteins and prevent their aggregation, and deliver unfolded protiens to HSP70 (2). HSP60 has primarily been known as a mitochondrial protein that is important for folding key proteins after import into the mitochondria (3). HSP60 is also present in the cytosol of many cells and is induced by stress, inflammatory and immune responses, autoantibodies correlated with Alzheimer's, coronary artery diseases, MS, and diabetes (4-7). Secretory and transmembrane proteins are synthesized on polysomes and translocate into the endoplasmic reticulum (ER) where they are often modified by the formation of disulfide bonds, amino-linked glycosylation and folding. The ER contains a pool of molecular chaperones including calnexin, BiP and protein disulfide isomerase (PDI). Calenxin is a calcium-binding protein embedded in the ER membrane that retains newly synthesized glycoproteins inside the ER to ensure proper folding and quality control (8,9). When protein folding is disturbed inside the ER, Bip synthesis is increased. Subsequently, BiP binds to misfolded proteins to prevent them from forming aggregates and assists them to refold properly (10). PDI catalyzes the formation and isomerization of disulfide bonds required to reach a proteins native state (11). Heat shock gene transcription is regulated by a familly of heat shock factors (HSFs), transcriptional activators that bind to heat shock response elements (HSEs) located upstream of all heat shock genes (12). During attenuation from the heat shock response, HSF1 is repressed by direct binding of HSP70, HSP40/Hdj-1 and HSF binding protein 1 (HSBP1) (13).

  1. Nollen, E.A. and Morimoto, R.I. (2002) J. Cell Sci. 115, 2809-2816.
  2. Fan, C.Y. et al. (2003) Cell Stress Chaperones 8, 309-316.
  3. Jindal, S. et al. (1989) Mol Cell Biol 9, 2279-83.
  4. Itoh, H. et al. (2002) Eur. J. Biochem. 269, 5931-5938.
  5. Gupta, S. and Knowlton, A.A. J. Cell Mol. Med. 9, 51-58.
  6. Deocaris, C.C. et al. (2006) Cell Stress Chaperones 11, 116-128.
  7. Lai, H.C. et al. (2007) Am. J. Physiol. Endocrinol. Metab. 292, E292-E297.
  8. Bergeron, J.J. et al. (1994) Trends Biochem. Sci. 19, 124-128.
  9. Williams, D.B. (2006) J. Cell Sci. 119, 615-623.
  10. Kohno, K. et al. (1993) Mol. Cell. Biol. 13, 877-890.
  11. Ellgaard, L. and Ruddock, L.W. (2005) EMBO Rep. 6, 28-32.
  12. Morimoto, R.I. (1998) Genes Dev. 12, 3788-3796.
  13. Satyal, S.H. et al. (1998) Genes Dev. 12, 1962-1974.

Application References

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Protocols

* Product-specific protocol.

Companion Products

For Research Use Only. Not For Use In Diagnostic Procedures.

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