Overview

  • Product name
  • Description
    Rabbit polyclonal to Haspin
  • Host species
    Rabbit
  • Tested applications
    Suitable for: WBmore details
  • Species reactivity

    Predicted to work with: Human
  • Immunogen

    Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Human Haspin.

    Read Abcam's proprietary immunogen policy (Peptide available as ab23532.)

  • Positive control
    • This antibody gave a positive signal in the following whole cell lysates: HeLa Expressing myc-Haspin (interphase); HeLa Expressing myc-Haspin (mitosis) Whole Cell Lysate.

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
  • Storage buffer
    Preservative: 0.02% Sodium Azide
    Constituents: 1% BSA, PBS, pH 7.4
  • Concentration information loading...
  • Purity
    Immunogen affinity purified
  • Clonality
    Polyclonal
  • Isotype
    IgG
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab21686 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 0.2 µg/ml. Detects a band of approximately 90 kDa (predicted molecular weight: 88 kDa).

Target

  • Function
    Required for normal alignment of chromosomes at metaphase. Phosphorylates histone H3 'Thr-3' during mitosis.
  • Tissue specificity
    Strongly expressed in testis. Also present in thymus and bone marrow and low levels observed in prostate, intestine, lung, spleen and lymph node. Expressed in fetal skin, liver, kidney and small intestine and also in proliferating but not non-proliferating cell lines.
  • Sequence similarities
    Belongs to the protein kinase superfamily. Ser/Thr protein kinase family. Haspin subfamily.
    Contains 1 protein kinase domain.
  • Post-translational
    modifications
    Autophosphorylated on both serine and threonine residues (By similarity). Phosphorylated during mitosis. Phosphorylated upon DNA damage, probably by ATM or ATR.
  • Cellular localization
    Nucleus.
  • Information by UniProt
  • Database links
  • Alternative names
    • germ cell associated 2 antibody
    • Germ cell-specific gene 2 protein antibody
    • GSG2 antibody
    • H-haspin antibody
    • Haploid germ cell-specific nuclear protein kinase antibody
    • HASP_HUMAN antibody
    • Serine/threonine-protein kinase haspin antibody
    see all

Images

  • All lanes : Anti-Haspin antibody (ab21686) at 1 µg/ml

    Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate at 10 µg
    Lane 2 : HeLa Expressing myc-Haspin (interphase) Whole Cell Lysate - Positive Control at 5 µl
    Lane 3 : HeLa Interphase Whole Cell Lysate - Negative Control at 10 µg
    Lane 4 : HeLa Expressing myc-Haspin (mitosis) Whole Cell Lysate - Positive Control at 5 µl
    Lane 5 : HeLa Mitosis Whole Cell Lysate - Negative Control at 10 µg

    Secondary
    All lanes : Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (HRP), pre-adsorbed at 1/5000 dilution

    Performed under reducing conditions.

    Predicted band size: 88 kDa
    Observed band size: 90 kDa
    why is the actual band size different from the predicted?
    Additional bands at: 23 kDa. We are unsure as to the identity of these extra bands.


    Exposure time: 4 minutes

References

This product has been referenced in:
  • Varier RA  et al. A phospho/methyl switch at histone H3 regulates TFIID association with mitotic chromosomes. EMBO J 29:3967-78 (2010). Read more (PubMed: 20953165) »
  • Markaki Y  et al. Phosphorylation of histone H3 at Thr3 is part of a combinatorial pattern that marks and configures mitotic chromatin. J Cell Sci 122:2809-19 (2009). WB ; Human, Mouse . Read more (PubMed: 19622635) »
See all 3 Publications for this product

Customer reviews and Q&As

1-2 of 2 Abreviews or Q&A

Question

DESCRIPTION OF THE PROBLEM No signal or weak signal SAMPLE Whole cell lysate of HEK293A, SHSY5Y, A549 treated with 50ng/ml of Nocodazole for 12 hours. PRIMARY ANTIBODY Concentration or dilution 1:1000 to 1:100 Diluent buffer 5% Skim milk in TTBS Incubation time 12-20 hours Incubation temperature: 4oC DETECTION METHOD ECL POSITIVE AND NEGATIVE CONTROLS USED GAPDH was nicely visualized with all the same steps and materials stated above. ANTIBODY STORAGE CONDITIONS -20C SAMPLE PREPARATION After twice washings with PBS, cells were harvested and the lysed with lysis buffer (20mM Tris (pH 7.5), 150mM NaCl, 1mM EDTA, 1mM EGTA, 1% Triton X-100, 2.5mM sodium pyrophosphate, 1mM beta-glycerophosphate, 10mM Na3VO4, 1µg/ml Leupeptin, 1ug/ml Aprotinin, 1mM PMSF). Reducing agent 5% beta-mercaptoethanol and 50mM DTT Boiling for ≥5 min? 5 minutes AMOUNT OF PROTEIN LOADED Up to 50ug ELECTROPHORESIS/GEL CONDITIONS SDS-PAGE, 12% TRANSFER AND BLOCKING CONDITIONS Lysates were separated by SDS-PAGE and transferred to nitrocellulose membrane at 100V for 60 minutes. Transfer buffer is composed of 25 mM Tris, 192 mM glycine and 20% methanol. Membrane was blocked with TTBS (150mM NaCl, 10mM Tris-HCl (pH 7.4), 0.05% Tween 20) containing 5% skim milk for 30min, and incubated with selected primary antibodies O/N Protein transfer verified -verified by Ponceau S Red staining SECONDARY ANTIBODY - Goat anti-Rabbit/blocking solution/1:1000/16 hours at 4oC/washed threetimes in TTBS for 10min with gentle rotation. HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 5 HAVE YOU RUN A "NO PRIMARY" CONTROL? No DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes WHAT STEPS HAVE YOU ALTERED? Dilution ratio was concentrated up to 1:100 and loading amounts of cell lysates was increased up to 100ug.

Read More
Answer

Thank you for contacting Abcam regarding ab21686. I am sorry that you have been experiencing difficulties with this antibody in WB. I have reviewed the protocol information provided and would like to make some suggestions to improve the results with this antibody. Have you tested the antibody with BSA as a blocking reagent? I know we have some data on our website to suggest that milk should also work, but we have tested and validated recent lots with 5% BSA as a blocking agent and antibody diluent. I would encourage you to test this as we have seen strong positive results with this protocol. I hope this information is helpful. Please do not hesitate to contact us if you have any additional questions or if these suggestions do not improve your results with this antibody.

Read More
Application
Western blot
Sample
Human Cell lysate - whole cell (HeLa)
Loading amount
17 µg
Specification
HeLa
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5%

Dr. Jonathan Higgins

Verified customer

Submitted Feb 12 2007

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

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