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Signal Transduction Metabolism Mitochondrial
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Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)

  • Datasheet
  • SDS
Reviews (1)Q&A (2)References (10)

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Western blot - Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)
  • Immunocytochemistry/ Immunofluorescence - Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)
  • Flow Cytometry - Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)
  • Western blot - Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)

Key features and details

  • Mouse monoclonal [3B7BH7] to ACADM/MCAD
  • Suitable for: WB, Flow Cyt, IHC-P, ICC/IF
  • Reacts with: Mouse, Rat, Human
  • Isotype: IgG1

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Alexa Fluor® 488 Alexa Fluor® 647

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Overview

  • Product name

    Anti-ACADM/MCAD antibody [3B7BH7]
    See all ACADM/MCAD primary antibodies
  • Description

    Mouse monoclonal [3B7BH7] to ACADM/MCAD
  • Host species

    Mouse
  • Tested applications

    Suitable for: WB, Flow Cyt, IHC-P, ICC/IFmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Recombinant full length protein. This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • HeLa cells, HL-60 cells, Human cerebellum, HepG2 cells, HeLa cells, H9C2 (rat cells), and MEF (mouse cells) lysates.
  • General notes

    MCAD (medium-chain acyl-CoA dehydrogenase) is an oxidoreductase enzyme of the mitochondrial fatty acid beta-oxidation pathway that is specific for acyl chain lengths of 4 to 16. It also utilizes the electron transfer flavoprotein (ETF) as electron acceptor that transfers the electrons to the main mitochondrial respiratory chain via ETF-ubiquinone oxidoreductase (ETF dehydrogenase).

    This antibody clone is manufactured by Abcam. If you require a custom buffer formulation or conjugation for your experiments, please contact orders@abcam.com.

    The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing this with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation. Please check that this product meets your needs before purchasing.

    If you have any questions, special requirements or concerns, please send us an inquiry and/or contact our Support team ahead of purchase. Recommended alternatives for this product can be found below, along with publications, customer reviews and Q&As

    Product was previously marketed under the MitoSciences sub-brand.

Properties

  • Form

    Liquid
  • Storage instructions

    Shipped at 4°C. Store at +4°C. Do Not Freeze.
  • Storage buffer

    pH: 7.5
    Preservative: 0.02% Sodium azide
    Constituent: 99.98% HEPES buffered saline
  • Concentration information loading...
  • Purity

    Ammonium Sulphate Precipitation
  • Purification notes

    Produced in vitro using hybridomas grown in serum-free medium, and then purified by biochemical fractionation.
  • Primary antibody notes

    MCAD (medium-chain acyl-CoA dehydrogenase) is an oxidoreductase enzyme of the mitochondrial fatty acid beta-oxidation pathway that is specific for acyl chain lengths of 4 to 16. It also utilizes the electron transfer flavoprotein (ETF) as electron acceptor that transfers the electrons to the main mitochondrial respiratory chain via ETF-ubiquinone oxidoreductase (ETF dehydrogenase).
  • Clonality

    Monoclonal
  • Clone number

    3B7BH7
  • Isotype

    IgG1
  • Light chain type

    kappa
  • Research areas

    • Signal Transduction
    • Metabolism
    • Mitochondrial
    • Signal Transduction
    • Metabolism
    • Lipid metabolism
    • Cardiovascular
    • Lipids / Lipoproteins
    • Fatty Acids
    • Metabolism
    • Cancer
    • Cancer Metabolism
    • Metabolic signaling pathway
    • Metabolism of lipids and lipoproteins
    • Metabolism
    • Pathways and Processes
    • Mitochondrial Metabolism
    • Mitochondrial markers
    • Metabolism
    • Pathways and Processes
    • Metabolic signaling pathways
    • Lipid and lipoprotein metabolism
    • Lipid metabolism
    • Metabolism
    • Types of disease
    • Cancer
    • Metabolism
    • Pathways and Processes
    • Redox metabolism
    • Fatty acid oxidation

Associated products

  • Alternative Versions

    • Alexa Fluor® 488 Anti-ACADM/MCAD antibody [3B7BH7] (ab198078)
    • Alexa Fluor® 647 Anti-ACADM/MCAD antibody [3B7BH7] (ab198099)
  • Compatible Secondaries

    • Goat Anti-Mouse IgG H&L (Alexa Fluor® 488) (ab150113)
    • Goat Anti-Mouse IgG H&L (HRP) (ab205719)
  • Conjugation kits

    • PE / R-Phycoerythrin Conjugation Kit - Lightning-Link® (ab102918)
    • APC Conjugation Kit - Lightning-Link® (ab201807)
  • Isotype control

    • Mouse IgG1, kappa monoclonal [15-6E10A7] - Isotype Control (ab170190)
  • Positive Controls

    • HeLa membrane extract lysate (ab29547)
  • Recombinant Protein

    • Recombinant Human ACADM/MCAD protein (ab99329)
  • Related Products

    • Fenofibrate, PPAR-alpha agonist (ab120832)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab110296 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 (1)
Use a concentration of 0.125 µg/ml. Predicted molecular weight: 47 kDa.
Flow Cyt
Use a concentration of 1 µg/ml.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody. 

IHC-P
1/1000. Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.
ICC/IF
Use a concentration of 1 µg/ml.
Notes
WB
Use a concentration of 0.125 µg/ml. Predicted molecular weight: 47 kDa.
Flow Cyt
Use a concentration of 1 µg/ml.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody. 

IHC-P
1/1000. Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.
ICC/IF
Use a concentration of 1 µg/ml.

Target

  • Function

    This enzyme is specific for acyl chain lengths of 4 to 16.
  • Pathway

    Lipid metabolism; mitochondrial fatty acid beta-oxidation.
  • Involvement in disease

    Defects in ACADM are the cause of acyl-CoA dehydrogenase medium-chain deficiency (ACADMD) [MIM:201450]. It is an autosomal recessive disease which causes fasting hypoglycemia, hepatic dysfunction, and encephalopathy, often resulting in death in infancy.
  • Sequence similarities

    Belongs to the acyl-CoA dehydrogenase family.
  • Cellular localization

    Mitochondrion matrix.
  • Target information above from: UniProt accession P11310 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 34 Human
    • Entrez Gene: 11364 Mouse
    • Entrez Gene: 24158 Rat
    • Omim: 607008 Human
    • SwissProt: P11310 Human
    • SwissProt: P45952 Mouse
    • SwissProt: P08503 Rat
    • Unigene: 445040 Human
    • Unigene: 10530 Mouse
    • Unigene: 6302 Rat
    see all
  • Alternative names

    • ACAD 1 antibody
    • ACAD1 antibody
    • Acadm antibody
    • ACADM_HUMAN antibody
    • Acyl coenzyme A dehydrogenase antibody
    • Acyl coenzyme A dehydrogenase C 4 to C 12 straight chain antibody
    • FLJ18227 antibody
    • FLJ93013 antibody
    • FLJ99884 antibody
    • MCAD antibody
    • MCADH antibody
    • Medium chain acyl CoA dehydrogenase antibody
    • Medium chain fatty acyl CoA dehydrogenase antibody
    • Medium chain specific acyl CoA dehydrogenase antibody
    • Medium chain specific acyl CoA dehydrogenase mitochondrial antibody
    • Medium-chain specific acyl-CoA dehydrogenase antibody
    • mitochondrial antibody
    see all

Images

  • Western blot - Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)
    Western blot - Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)
    All lanes : Anti-ACADM/MCAD antibody [3B7BH7] (ab110296) at 0.125 µg/ml

    Lane 1 : HepG2 cell lysates
    Lane 2 : HeLa cell lysates
    Lane 3 : H9C2 (rat cells) lysates
    Lane 4 : MEF (mouse cells) lysates

    Lysates/proteins at 15 µg per lane.

    Predicted band size: 47 kDa

  • Immunocytochemistry/ Immunofluorescence - Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)
    Immunocytochemistry/ Immunofluorescence - Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)
    Immunocytochemistry image of ab110296 stained Human HeLa cells. The cells were paraformaldehyde fixed (4%, 20 minutes) and Triton X-100 permeabilized (0.1%, 15 minutes). The cells were incubated with ab110296 (1 µg/ml) for 2 hours at room temperature or over night at 4°C. The secondary antibody was (green) Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 dilution for 1 hour. 10% Goat serum was used as the blocking agent for all blocking steps. DAPI was used to stain the cell nuclei (blue). Target protein locates mainly in mitochondria.
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)

    ACADM/MCAD immunohistochemistry in Human cerebellum visualized with ab110296 at 1/1000. MCAD immunoactivity is most intense in neuronal cell bodies, most notably in the large Purkinje cells. Note the distinctive subcellular localization of MCAD immunoreactivity in the Purkinje cell bodies.

  • Flow Cytometry - Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)
    Flow Cytometry - Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)
    HL-60 cells were stained with 1 µg/mL ab110296 (blue) or an equal amount of an isotype control antibody (red) and analyzed by flow cytometry.
  • Western blot - Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)
    Western blot - Anti-ACADM/MCAD antibody [3B7BH7] (ab110296)

    HL-60 cells were incubated at 37&degC for 24h with vehicle control (0 &microM) and different concentrations of fenofibrate (ab120832). Increased expression of ACADM/MCAD in HL-60 cells correlates with an increase in fenofibrate concentration, as described in literature.

    Whole cell lysates were prepared with RIPA buffer (containing protease inhibitors and sodium orthovanadate), 10&microg of each were loaded on the gel and the WB was run under reducing conditions. After transfer the membrane was blocked for an hour using 5% BSA before being incubated with ab110296 at 1 &microg/ml and ab9484 at 1 &microg/ml overnight at 4°C. Antibody binding was detected using an anti-mouse antibody conjugated to HRP (ab97040) at 1/10000 dilution and visualised using ECL development solution.

     

Protocols

  • Mouse on Mouse staining protocol

Click here to view the general protocols

Datasheets and documents

  • SDS download

  • Datasheet download

    Download

References (10)

Publishing research using ab110296? Please let us know so that we can cite the reference in this datasheet.

ab110296 has been referenced in 10 publications.

  • Zhao J  et al. Mitochondrial Functions Are Compromised in CD4 T Cells From ART-Controlled PLHIV. Front Immunol 12:658420 (2021). PubMed: 34017335
  • Gao K  et al. Qishen granules exerts cardioprotective effects on rats with heart failure via regulating fatty acid and glucose metabolism. Chin Med 15:21 (2020). PubMed: 32158496
  • Zhou C  et al. Gadofullerene inhibits the degradation of apolipoprotein B100 and boosts triglyceride transport for reversing hepatic steatosis. Sci Adv 6:N/A (2020). PubMed: 32917715
  • Lim SC  et al. Loss of the Mitochondrial Fatty Acid ß-Oxidation Protein Medium-Chain Acyl-Coenzyme A Dehydrogenase Disrupts Oxidative Phosphorylation Protein Complex Stability and Function. Sci Rep 8:153 (2018). WB ; Human . PubMed: 29317722
  • Shaltouki A  et al. Alpha-synuclein delays mitophagy and targeting Miro rescues neuron loss in Parkinson's models. Acta Neuropathol 136:607-620 (2018). PubMed: 29923074
  • Cornelius N  et al. Cellular consequences of oxidative stress in riboflavin responsive multiple acyl-CoA dehydrogenation deficiency patient fibroblasts. Hum Mol Genet N/A:N/A (2014). PubMed: 24698980
  • Tomilov A  et al. Shc depletion stimulates brown fat activity in vivo and in vitro. Aging Cell 13:1049-58 (2014). Mouse . PubMed: 25257068
  • Peek CB  et al. Circadian clock NAD+ cycle drives mitochondrial oxidative metabolism in mice. Science 342:1243417 (2013). PubMed: 24051248
  • Tucci S  et al. Tissue-Specific Strategies of the Very-Long Chain Acyl-CoA Dehydrogenase-Deficient (VLCAD(-/-)) Mouse to Compensate a Defective Fatty Acid ß-Oxidation. PLoS One 7:e45429 (2012). WB ; Mouse . PubMed: 23024820
  • Frier BC  et al. Interactions between the consumption of a high-fat diet and fasting in the regulation of fatty acid oxidation enzyme gene expression: an evaluation of potential mechanisms. Am J Physiol Regul Integr Comp Physiol 300:R212-21 (2011). PubMed: 21084676

Customer reviews and Q&As

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1-3 of 3 Abreviews or Q&A

Western blot abreview for Anti-ACADM/MCAD antibody [3B7BH7]

Good
Abreviews
Abreviews
abreview image
Application
Western blot
Sample
Rat Tissue lysate - whole (Heart tissue lysate)
Gel Running Conditions
Reduced Denaturing (10%)
Loading amount
30 µg
Specification
Heart tissue lysate
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 30°C
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The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.

Dr. SAIFUDEEN ISMAEL

Verified customer

Submitted Feb 15 2013

Question

in Ihrem Kit handelt es sich um einen Sandwich-Assay. Es kommen zwei Antikörper zum Einsatz. Welcher der beiden ist der AB110296, der immobilisierte oder der im nächsten Schritt zur Reaktion dazugegebene AK? Gibt es Angaben zu diesem anderen Antikörper?
Die in Ihrem Kit angegebene Mutante c.985A>G führte mit den in der Reaktion verwendeten AK zu keiner Reaktion. Sie ziehen aus dieser Tatsache den Schluss, dass die Mutante instabil ist und deshalb entweder nicht gebildet oder bereits wieder degradiert wurde. Gibt es dazu weiterführende Literatur oder zumindest einen Western Blot, der bestätigt, dass das Protein wirklich nicht da ist? Ist es nicht wahrscheinlicher, dass die Mutante einer der beiden AK nicht binden kann auf Grund einer Konformationsänderung und es deshalb zu keiner Farbreaktion kommt?
Wir haben einen ähnlichen Versuch mit zwei anderen Mutanten (liegen beide vor bzw. weit vor der 985) durchgeführt. Im Western Blot sind beide Nachweisbar unter Verwendung von AB110296, zeigen eine geringe Enzym-Aktivität, bringen aber ebenfalls keine Farbreaktion mit dem ACADM Elisa Kit, während sich das WT-Protein problemlos anfärben lässt.
Mit der Bitte um Antwort und freundlichem Gruß

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Abcam community

Verified customer

Asked on Aug 06 2012

Answer

Vielen Dank für Ihre Antwort.

Leider ist mir bei meiner letzten Antwort ein Fehler unterlaufen und ich möchte diesen hiermit korrigieren:

ab110296 wird nicht im Kit ACADM ELISA Kit (ab129734) verwendet.. Es wird ein Antikörper verwendet, der nicht einzeln erhältlich ist. Bitte entschuldigen Sie vielmals diese Fehlinformation.

Bezüglich der anderen Fragen habe ich unsere Spezialisten bei MitoSciences gefragt. Ich habe ihnen die Antwort direkt unten rein kopiert:

"We purchased the cell line with the K304E (984A>G in ACADM gene) from the Coriell repository. The cell line is homozygous for 984A>G . The mutation in this cell line was found with a decrease in MCAD activity (12% from control) by the submitter, but there is no specific reference for this particular cell line in the coriell website.

http://ccr.coriell.org/Sections/Search/Sample_Detail.aspx?Ref=GM07844&PgId=166



However, it is well known in the literature that the K304E aa change (homozygous or compound heterozygous) induces MCAD deficiency (measured by enzyme activity). References below. Many of these studies were done before there were commercially available MCAD antibodies. So the WB data does not exist. Therefore we tested the above cell line from Coriell with our antibody 3B7BH7 and found that MCAD protein levels are significantly decreased (both by WB and by ICC). The customer can look at the data in the ICE booklet as mentioned in my previous email. We don’t have WB data with the antibody used in the kit showing control vs deficient, because the antibody is WB negative.

A good place to look at mutations known to induce MCAD deficiency is OMIM.

http://omim.org/entry/201450



http://omim.org/entry/201450#reference32 reported the findings in a prospective neonatal screening program in Pennsylvania using tandem mass spectrometry. From the first 80,371 newborns screened, they found 9 babies with MCAD (1/8930), plus 2 additional newborns, screened because of a previously known family history. Molecular analysis showed that 56% of the patients were compound heterozygotes for the 985A-G mutation (K304E; http://omim.org/entry/607008#0001), commonly referred to as G985, and a second mutation.


In 9 patients with MCAD deficiency, http://omim.org/entry/201450#reference16 found a 985A-G transition which resulted in a lys304-to-glu substitution (K304E; http://omim.org/entry/607008#0001) in the mature protein. These patients were unrelated, suggesting a high incidence of this abnormality among Caucasian patients. The change was not found in 20 healthy Caucasian and 6 healthy Japanese subjects. http://omim.org/entry/201450#reference16 found this point mutation in 31 of 34 (91%) mutant MCAD alleles. http://omim.org/entry/201450#reference31 estimated that 90% of MCAD cases involve the same mutation.

http://omim.org/entry/201450#reference33 characterized the molecular defect in 4 patients with mild MCAD deficiency. In routine neonatal screening on the fifth day of life, they had been found to have abnormal acylcarnitine profiles indicative of MCAD deficiency. Two were of German origin and the other 2 were born to different consanguineous Turkish parents. In all 4, the clinical course and routine laboratory investigations up to the age of 6 months were unremarkable. Enzyme studies showed residual MCAD activities between those with classic MCAD deficiency and heterozygotes. In 2 cases, ACADM gene analysis revealed compound heterozygosity for the common K304E mutation (http://omim.org/entry/607008#0001) and the 199T-C mutation (Y42H; http://omim.org/entry/607008#0011), which they designated Y67H. In the 2 children of consanguineous parents, homozygosity was found for the gly267-to-arg mutation (G267R; http://omim.org/entry/607008#0003) and the S220L mutation (http://omim.org/entry/607008#0012), respectively. As in other metabolic disorders, the distinction between 'normal' and 'disease' in MCAD deficiency is blurred into a spectrum of enzyme deficiency states caused by different mutations in the ACADM gene potentially influenced by factors affecting intracellular protein processing."



Ich hoffe, diese Information ist hilfreich und wird Ihnen Ihre Bedenken bezüglich des Kits nehmen.

Read More

Abcam Scientific Support

Answered on Aug 06 2012

Question

Inquiry: Sehr geehrte Damen und Herren, für meine Experimente nutze ich zur Zeit neben dem monoklonalen MCAD-Antikörper (MS726) auch das MCAD ELISA Kit (ab129734). Für die Auswertung meiner Versuche wäre es sehr wichtig für mich, die Sequenz zu wissen, an welche der Antikörper bindet und wo diese sich befindet, da anderweitig eine Auswertung kaum möglich ist. Ich würde mich deswegen sehr freuen, wenn Sie mir diese Sequenz mitteilen würden. Desweiteren würde ich gerne wissen, ob der monoklonsle AK (siehe oben), den wir einzeln bestellt haben, der AK ist, der auch im Kit zur Anwendung kommt. Im Kit ist das leider nicht ohne Weiteres nachzuvollziehen. Über Ihre Hilfe würde ich mich wirklich sehr freuen! Mit besten Grüßen aus der

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Abcam community

Verified customer

Asked on Jul 30 2012

Answer

Vielen Dank für Ihre Anfrage.

Das Immunogen für ab110296 war das Protein voller Länge. Das Epitop, das der Antikörper bindet ist leider nicht näher kartiert worden und wir kennen es deshalb nicht.

Ich freue mich Ihnen bestätigen zu können, dass ab110296 im Kit ACADM ELISA Kit (ab129734) verwendet wird.

Es tut mir leid, dass ich Ihre Fragen diesmal nicht vollständig beantworten konnte und hoffe, dass diese Information dennoch hilfreich ist.

Read More

Abcam Scientific Support

Answered on Jul 30 2012

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