Overview

  • Product name
  • Description
    Goat polyclonal to Hsp22
  • Host species
    Goat
  • Tested applications
    Suitable for: WB, ICCmore details
  • Species reactivity
    Reacts with: Human
  • Immunogen

    Synthetic peptide:

    NELPQDSQEVTCT

    , corresponding to C terminal amino acids 184-196 of Human Hsp22.

  • Positive control
    • Recombinant Human Hsp22 protein (ab119152) can be used as a positive control in WB. Human Muscle lysate.
  • General notes
    Principal Names - H11; E2IG1; HSP22; HspB8; protein kinase H11; small stress protein-like protein HSP22 Official Gene Symbol - H11 (interim symbol) GenBank Accession Number – NP_055180 LocusLink ID - 26353 (human) Gene Ontology terms - protein serine/threonine kinase activity; heat shock protein activity; biological_process unknown; cellular_component unknown; transferase activity.

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Upon delivery aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.
  • Storage buffer
    Preservative: 0.02% Sodium Azide
    Constituents: 0.5% BSA, 5mg/ml Tris, pH 7.3
  • Concentration information loading...
  • Purity
    Immunogen affinity purified
  • Purification notes
    Purified from goat serum by ammonium sulphate precipitation followed by antigen affinity chromatography using the immunizing peptide.
  • Clonality
    Polyclonal
  • Isotype
    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab4149 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
WB Use a concentration of 0.5 - 1 µg/ml. Detects a band of approximately 20-25 kDa (predicted molecular weight: 22 kDa).Can be blocked with Human Hsp22 peptide (ab22992).

Approx 20-25 kDa band seen in Human Muscle lysate [Predicted MW of approx. 22 kDa according to NP_055180].

ICC Use at an assay dependent concentration.

Target

  • Function
    Displays temperature-dependent chaperone activity.
  • Tissue specificity
    Predominantly expressed in skeletal muscle and heart.
  • Involvement in disease
    Neuronopathy, distal hereditary motor, 2A
    Charcot-Marie-Tooth disease 2L
  • Sequence similarities
    Belongs to the small heat shock protein (HSP20) family.
  • Cellular localization
    Cytoplasm. Nucleus. Translocates to nuclear foci during heat shock.
  • Information by UniProt
  • Database links
  • Alternative names
    • Alpha crystallin C chain antibody
    • Alpha-crystallin C chain antibody
    • Charcot Marie Tooth disease axonal type 2L antibody
    • Charcot Marie Tooth disease spinal antibody
    • CMT2L antibody
    • CRYAC antibody
    • DHMN 2 antibody
    • DHMN2 antibody
    • E2 induced gene 1 protein antibody
    • E2-induced gene 1 protein antibody
    • E2IG1 antibody
    • H11 antibody
    • Heat shock 22kDa protein 8 antibody
    • Heat shock 27kDa protein 8 antibody
    • Heat shock protein 22 antibody
    • Heat shock protein beta 8 antibody
    • Heat shock protein beta-8 antibody
    • Hereditary motor neuropathy distal antibody
    • HMN 2 antibody
    • HMN2 antibody
    • HMN2A antibody
    • HSB8 antibody
    • HSPB 8 antibody
    • HspB8 antibody
    • HSPB8_HUMAN antibody
    • OTTHUMP00000239768 antibody
    • Protein kinase H11 antibody
    • Small stress protein like protein HSP22 antibody
    • Small stress protein-like protein HSP22 antibody
    • Spinal muscular atrophy distal adult autosomal dominant antibody
    see all

Images

  • ab4149 staining (1µg/ml) of Human Muscle lysate (RIPA buffer, 30µg total protein per lane).  Primary incubated for 1 hour.  Detected by western blot using chemiluminescence. ab4149 staining (1µg/ml) of Human Muscle lysate (RIPA buffer, 30µg total protein per lane). Primary incubated for 1 hour. Detected by western blot using chemiluminescence.

References

This product has been referenced in:
  • Sarparanta J  et al. Mutations affecting the cytoplasmic functions of the co-chaperone DNAJB6 cause limb-girdle muscular dystrophy. Nat Genet 44:450-5 (2012). Read more (PubMed: 22366786) »
  • Laure L  et al. Cardiac H11 kinase/Hsp22 stimulates oxidative phosphorylation and modulates mitochondrial reactive oxygen species production: Involvement of a nitric oxide-dependent mechanism. Free Radic Biol Med 52:2168-2176 (2012). WB . Read more (PubMed: 22542467) »
See all 3 Publications for this product

Customer reviews and Q&As

1-5 of 5 Abreviews or Q&A

Abcam has not validated the combination of species/application used in this Abreview.
Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (Embryonic Heart)
Permeabilization
Yes - Tween-20
Specification
Embryonic Heart
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Fixative
Paraformaldehyde

Abcam user community

Verified customer

Submitted Aug 22 2016

Abcam has not validated the combination of species/application used in this Abreview.
Application
Western blot
Sample
Mouse Tissue lysate - whole (Heart)
Gel Running Conditions
Reduced Denaturing (4-12%)
Loading amount
20 µg
Specification
Heart
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Aug 21 2016

Abreviews
Application
Western blot
Sample
Human Cell lysate - whole cell (Hela)
Loading amount
60 µg
Specification
Hela
Gel Running Conditions
Reduced Denaturing
Blocking step
Milk as blocking agent for 30 minute(s) · Concentration: 2% · Temperature: 21°C

Mr. Jörg Hoehfeld

Verified customer

Submitted Jun 27 2007

Answer

Thank you for taking the time to contact me regarding your experiences using this product, I am sorry to hear that you have been experiencing problems using this product in Western blotting. The working dilution (assuming a start conc. of 0.5mg/ml) ranges between 1/250 and 1/500, it may be necessary to go down to 1/250 (0.5ug/ml) to achieve a signal. Having looked at your previous order for this product back in 2003, it would appear that the product may have been at a different concentration (same mass), which is perhaps where 1:500-1:1000 dilutions are derived from? I would advise using the Ab at 1/300. If after increasing the concentration of the primary Ab you are still having problems using this product then please do not hesitate to get back in touch with me. All our customer feedback, including complaints are monitored weekly by our in house technical support team. If a product is at fault the technical support team will consider removing the product from our catalogue in order to avoid future customer inconvenience.

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Question

Thank you for your reply. I have attached a soft copy of the blot together with the markers. Please refer to below for the answers to your questions. 1. Please describe the problem (high background, wrong band size, more bands, no band etc). No band, presence of nonspecific bindings. 2. On what material are you testing the antibody in WB? • Species? Human SH-SY5Y cells • Cell extract/ Nuclear extract? Cell protein lysate • Purified protein? • Recombinant protein? 3. How much protein did you load? 15 ug • How did you prepare the lysate for the analysis (protease inhibitors etc)? Cells were lysed in RIPA buffer (10 mM Tris-HCl, pH 7.4, 150 mM NaCl, 1 mM EDTA, 1% Nonidet-P 40, 0.5% sodium deoxycholate, and 0.1% SDS) containing protease inhibitor cocktail tablet (Roche Diagnostics GmbH, Mannheim, Germany) and the supernatant collected after centrifugation at 13000 rpm for 10min. Equal amounts of protein (15ug) were separated by SDS-PAGE, transferred to polyvinyl difluoride (PVDF) membranes and probed with the primary antibody. • Did you heat the samples? Yes heated after addition of sample loading buffer containing 20% beta-mercaptoethanol, 5min, 100oC. 4. Primary Antibody • Specification (in which species was it raised against)? Goat • At what dilution(s) have you tested this antibody? 1 in 500 • Incubation time, wash step? overnight, 4oC, shaking. 5. Secondary Antibody • Specification (in which species was it raised against)? Donkey anti goat • At what dilution(s) have you tested this antibody? 1 in 10000 • Incubation time, wash step? 1 h, room temperature; wash 4 times, 15 min each in TBST • Do you know whether the problems you are experiencing come from the secondary? Unlikely as it gives good results with other goat antibodies. 6. What detection method are you using? Chemiluminescence 7. Background bands • Have you eliminated the possibility that any background bands could be due to the secondary antibody? (Run a “No primary” control) • Is the blocking step sufficient? (We recommend blocking the membrane by adding 20 ml of blocking buffer (5% non-fat dry milk, 0.1% Tween-20 in TBS). Incubate for 2 h at room temperature or overnight at 4°C with agitation) Blocking using 50% goat serum diluted in TBST, RT 1h. • Are your washing steps sufficiently stringent? (Multiple short washes are more effective than fewer longer wash steps) • At what size are the bands migrating? Could they be degradation products of your target? • Please provide an image of your blot (as an e-mail attachment, a faxed image is not sufficient) 8. Optimization attempts • How many times have you tried the Western? Several times against transfected mouse HSP22 but perhaps it does not recognize the mouse protein although there are only 2 different amino acid residues. The last time was against human HSP22 which should be overexpressed by our treatment of SY5Y cells. • Do you obtain the same results every time e.g. are background bands always in the same place? • What steps have you altered? 9. Did you apply positive and negative controls along with the samples? Please specify. We have no positive controls.

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Answer

Thank you for submitting the completed Western Blot Questionnaire Form. We are very sorry to hear that you are having problem with this antibody. We would like to make the following comments/suggestions: 1. This antibody has been tested and characterised on Human Muscle lysate. As you probably know Small stress protein-like protein HSP22 is predominantly expressed in skeletal muscle and in heart. Since you are using a human neuroblastoma cell line (SH-SY5Y) which we have never tested; we would strongly suggest running a positive control along with your samples. 2. You may also need to increase the amount of loaded material onto the gel. 15 ug protein is a bit too low, try to load at least 25-30 ug total prptein per lane. We hope this will help.

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Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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