Recombinant
RabMAb

Recombinant Anti-Hemoglobin subunit alpha antibody [EPR3608] (ab92492)

Overview

  • Product name

    Anti-Hemoglobin subunit alpha antibody [EPR3608]
    See all Hemoglobin subunit alpha primary antibodies
  • Description

    Rabbit monoclonal [EPR3608] to Hemoglobin subunit alpha
  • Host species

    Rabbit
  • Specificity

    This antibody gave a positive signal in Western blotting of rat brain, heart, kidney and spleen tissue. This antibody works in human, mouse and rat in IHC.
  • Tested applications

    Suitable for: ELISA, WB, IHC-P, Flow Cytmore details
    Unsuitable for: IP
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide corresponding to Human Hemoglobin subunit alpha aa 50-150.
    (Peptide available as ab206105)

  • Positive control

    • K562 cell lysate, human heart lysate, human spleen tissue, human stomach cancer tissue, human lung tissue, human placenta, rat heart lysate, mouse spleen lysate and mouse cerebral cortex.
  • General notes

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents.

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab92492 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
ELISA Use at an assay dependent concentration.
WB 1/1000 - 1/2000. Detects a band of approximately 14 kDa (predicted molecular weight: 15 kDa).Can be blocked with Hemoglobin subunit alpha peptide (ab206105).
IHC-P 1/100 - 1/500. Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.

See IHC antigen retrieval protocols.

Flow Cyt 1/100.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

  • Application notes
    Is unsuitable for IP.
  • Target

    • Function

      Involved in oxygen transport from the lung to the various peripheral tissues.
    • Tissue specificity

      Red blood cells.
    • Involvement in disease

      Heinz body anemias
      Alpha-thalassemia
      Alpha(0)-thalassemia is associated with non-immune hydrops fetalis, a generalized edema of the fetus with fluid accumulation in the body cavities due to non-immune causes. Non-immune hydrops fetalis is not a diagnosis in itself but a symptom, a feature of many genetic disorders, and the end-stage of a wide variety of disorders.
      Hemoglobin H disease
    • Sequence similarities

      Belongs to the globin family.
    • Post-translational
      modifications

      The initiator Met is not cleaved in variant Thionville and is acetylated.
    • Information by UniProt
    • Database links

    • Alternative names

      • Alpha 1 globin antibody
      • Alpha globin antibody
      • Alpha one globin antibody
      • Alpha-globin antibody
      • HBA_HUMAN antibody
      • HBA1 antibody
      • HBA2 antibody
      • Hemoglobin alpha 1 antibody
      • Hemoglobin alpha 1 chain antibody
      • Hemoglobin alpha 1 globin chain antibody
      • Hemoglobin alpha 2 antibody
      • Hemoglobin alpha chain antibody
      • Hemoglobin subunit alpha antibody
      • MGC126895 antibody
      • MGC126897 antibody
      see all

    Images

    • All lanes : Anti-Hemoglobin subunit alpha antibody [EPR3608] (ab92492) at 1/2000 dilution (purified)

      Lane 1 : Human heart lysate
      Lane 2 : Rat heart lysate

      Lysates/proteins at 15 µg per lane.

      Secondary
      All lanes : Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/10000 dilution

      Predicted band size: 15 kDa
      Observed band size: 14 kDa
      why is the actual band size different from the predicted?



      Blocking and diluting buffer: 5% NFDM/TBST.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human placenta tissue labeling Hemoglobin subunit alpha antibody with purified ab92492 at 1/500 dilution (1.4 μg/ml). Heat mediated antigen retrieval was performed using EDTA Buffer, PH9. ab97051, a HRP-conjugated goat anti-rabbit IgG H&L was used as the secondary antibody at 1/500 dilution. Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat kidney tissue labeling Hemoglobin subunit alpha antibody with purified ab92492 at 1/500 dilution (1.4 μg/ml). Heat mediated antigen retrieval was performed using EDTA Buffer, PH9. ab97051, a HRP-conjugated goat anti-rabbit IgG H&L was used as the secondary antibody at 1/500 dilution. Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

    • Anti-Hemoglobin subunit alpha antibody [EPR3608] (ab92492) at 1/10000 dilution (purified) + Mouse spleen lysate at 15 µg

      Secondary
      Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/10000 dilution

      Predicted band size: 15 kDa
      Observed band size: 14 kDa why is the actual band size different from the predicted?



      Blocking and diluting buffer and concentration: 5% NFDM /TBST. 

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse cerebral cortex tissue labeling Hemoglobin subunit alpha antibody with purified ab92492 at 1/500 dilution (1.4 μg/ml). Heat mediated antigen retrieval was performed using EDTA Buffer, PH9. ab97051, a HRP-conjugated goat anti-rabbit IgG H&L was used as the secondary antibody at 1/500 dilution. Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

    • All lanes : Anti-Hemoglobin subunit alpha antibody [EPR3608] (ab92492) at 1/1000 dilution (unpurified)

      Lane 1 : K562 cell lysate
      Lane 2 : Human heart lysate

      Lysates/proteins at 10 µg per lane.

      Secondary
      All lanes : HRP labelled goat anti-rabbit at 1/2000 dilution

      Predicted band size: 15 kDa
      Observed band size: 14 kDa why is the actual band size different from the predicted?

    • Immunohistochemical analysis of paraffin embedded human placenta tissue labeling hemoglobin subunit alpha with unpurified ab92492 at 0.051 µg/ml (1:40000 dilution).

      Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.

    • Immunohistochemical analysis of paraffin embedded human stomach cancer tissue labeling hemoglobin subunit alpha with unpurified ab92492 at 0.051 µg/ml (1:40000 dilution).

      Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.

    • Immunohistochemical analysis of paraffin embedded human placenta tissue labeling hemoglobin subunit alpha with unpurified ab92492 at 0.051 µg/ml (1:40000 dilution).

      Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.

    References

    This product has been referenced in:

    • Kamnaksh A  et al. Harmonization of pipeline for preclinical multicenter plasma protein and miRNA biomarker discovery in a rat model of post-traumatic epileptogenesis. Epilepsy Res 149:92-101 (2019). Read more (PubMed: 30553097) »
    • Turtle JD  et al. Engaging pain fibers after a spinal cord injury fosters hemorrhage and expands the area of secondary injury. Exp Neurol 311:115-124 (2019). Read more (PubMed: 30268767) »
    See all 11 Publications for this product

    Customer reviews and Q&As

    1-9 of 9 Abreviews or Q&A

    Answer

    I have processed your request to receive ab109444 as a replacement for ab92492. I have also requested that you receive a new vial of secondary antibody and a new HRP substrate kit as well. I do not have an estimated time of arrival yet, but I will keep you updated when I find out more information.

    Read More

    Question
    Answer

    Vielen Dank für Ihren Anruf.

    Es tut mir leid, dass Sie Probleme mit diesem Antikörper hatten. Wie am Telefon besprochen habe ich eine kostenlose Ersatzlieferung für Sie in Auftrag gegeben. Sie hat die Referenznummer 123456und sollteMorgen bei Ihnen ankommen.

    Ich wünsche Ihnen viel Erfolg für Ihr Projekt. Bitte zögern Sie nicht, sich wieder bei uns zu melden, falls Sie weitere Fragen haben.

    Read More

    Answer

    Thank you for contacting us. I am sorry to hear that ab92492 is producing unsatisfactory results in WB. All of our Hemoglobin subunit alpha antibodies have been validated for use on endogenous proteins, but have not been tested on purified hemoglobin protein. In response to your questions:

    1.) This was performed as a reduced and denatured WB using a Laemmli Sample Buffer containing SDS and beta-mercaptoethanol.

    2.) The antibody was validated using 5% milk for blocking and 5% BSA for the antibody diluent. We do not currently have any feedback to suggest that this antibody performs better with one blocking agent over another.

    3.) The wash and dilution buffers were TBST containing 0.1% Tween-20.

    4.) We would expect this antibody to produce good results using any standard WB protocol, but if you could send me the details of your protocol and a description of the problem (no signal, high background, etc), I will be happy to make more specific recommendations to help optimize your results.

    5.) The testing was performed using a 1:1000 dilution for the primary antibody with a 1:2000 dilution of the secondary antibody. Rabbit monoclonal antibodies are well known for having very high binding affinities, however, so it is possible that the antibody may be suitable for use at a lower concentration.

    I hope this helps, please let me know if you need any additional information or assistance.

    Read More

    Question
    Answer

    Thank you for sending that protocol information.

    If I have read the information correctly, then there are a few protocol variations that may work:

    1 - Did the customer only try a 1/5000 primary antibody dilution? If so, I strongly suggest that they try range of dilutions from 1/500-1/2000, to see if they can find the concentration that will work for them.

    2 - According to the protocol, they loaded 5ug of protein on the gel. Did they try increasing the amount of protein, to say 20ug?

    3 - How long did they incubate the primary antibody for? We recommend overnight at 4C.

    4 - Did they try using 5% milk as a blocking reagent instead of BSA?

    If the customer has not tried the protocol variations above, then I would strongly suggest that they do. If they have already tried those variations, then I would be happy to process a refund for this antibody, as it is not working as we guarantee.

    I look forward to your reply.

    Read More

    Answer

    Thank you for your interest in Abcam. We continually add antibodies and other protein detection products to our catalogue but we do not offer custom-made antibodies. However, we are happy to pass on the contact details of the following companies that do offer this service: United States Pocono Rabbit Farm and Laboratory (PRF&L) PRF&L PO Box 240 Canadensis, PA 18325 Tel: 800-622-6381 - Toll Free in the United States & Canada Tel: 570-595-7108 - Worldwide Fax: 570-595-9277 Website: www.prfal.com Maine Biotechnology Services, Inc. 1037R Forest Ave. Portland Maine 04103 Tel: 800-925-9476 or 207-797-5454 Fax: 207-797-5595 Website: www.mainebiotechnology.com Cocalico Biologicals, Inc. PO Box 265 449 Stevens Road Reamstown, PA 17567-0265 Tel: 717-336-1990 Fax: 717-336-1993 Website: www.cocalicobiologicals.com United Kingdom Charles River Laboratories International, Inc. Tel: +44 (0) 1843 823575 Website: www.criver.com I hope this information is helpful to you. Please do not hesitate to contact us if you have any questions.  

    Read More

    Answer

    I have talked to my colleagues and I will be able to send a new secondary antibody aswell as a new ECL kit with the replacement antibody. I will send the new products as soon as*****decides which antibody he would like to receive, either ab109444 or ab102758.

    Read More

    Answer

    Gracias por contactarnos de nuevo y enviarnos más información acerca del protocolo seguido. -¿Habéis llevado a cabo un control no primario? Esto es, incubar directamente el secundario y observar si existe tinción sin necesidad de haber añadido el primario. -Mencionas que el secundario es Rabbit biotinilado ¿quieres decir que la especie de la que procede el secundario es rabbit? ¿Contra qué especie está dirigido? ¿Habéis probado otras diluciones del secundario? Quizás obtendríais mejores resultados con diluciones menos concentradas. -Para evitar la tinción inespecífica y con mucho background que apuntáis, es imprescindible bloquear la biotina endógena del tejido, pues es la principal fuente de señal de fondo. -Nosotros normalmente llevamos a cabo el proceso de recuperación de la proteína a diferentes tiempos (5, 10, 15, 20, 25 y 30 minutos). Es posible que el método utilizado sea demasiado agresivo, lo cual puede haber afectado a la morfología de la sección, dando lugar a una tinción inespecífica. Si pudierais enviarnos una imagen de cualquiera de los tejidos teñidos con el anticuerpo, nos ayudaría mucho a entender mejor el problema y facilitaros soluciones. En caso de que estas sugerencias no ayuden a obtener mejores resultados, por favor no dudéis en contactarnos de nuevo, estaremos encantados de seguir ayudándoos.

    Read More
    Application
    Immunocytochemistry
    Sample
    Mouse Cultured Cells (Erythroleukemia cell line)
    Specification
    Erythroleukemia cell line
    Fixative
    None
    Permeabilization
    No

    Abcam user community

    Verified customer

    Submitted Apr 21 2011

    Application
    ELISA
    Sample
    Human Recombinant protein (Recombinant HBA1)
    Specification
    Recombinant HBA1
    Blocking step
    BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
    Type
    Indirect

    Abcam user community

    Verified customer

    Submitted Mar 07 2011

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