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Validated using a knockout cell lineRecombinantRabMAb

Recombinant Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)

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Western blot - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
  • Western blot - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
  • Western blot - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
  • Western blot - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
  • Immunohistochemistry (Frozen sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
  • Immunoprecipitation - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
  • Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR20535] to Alpha-synuclein - BSA and Azide free
  • Suitable for: WB, IHC-P, IHC-Fr, IP
  • Knockout validated
  • Reacts with: Mouse, Rat, Human, Recombinant fragment

Conjugates logo Related conjugates and formulations

Unconjugated

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Conjugation
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HRP Conjugation Kit - Lightning-Link® (ab102890)
Protein
Product image
Recombinant Human Alpha-synuclein protein aggregate (Active) (ab218819)
Knockout
Product image
Human SNCA (Alpha-synuclein) knockout HEK-293T cell line (ab255433)

View more associated products

Overview

  • Product name

    Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free
    See all Alpha-synuclein primary antibodies
  • Description

    Rabbit monoclonal [EPR20535] to Alpha-synuclein - BSA and Azide free
  • Host species

    Rabbit
  • Tested applications

    Suitable for: WB, IHC-P, IHC-Fr, IPmore details
  • Species reactivity

    Reacts with: Mouse, Rat, Human, Recombinant fragment
  • Immunogen

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

  • Positive control

    • IHC-P: Human cerebral cortex tissue.
  • General notes

    ab225866 is the carrier-free version of ab212184.

    Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.

    This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.

    Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

    This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.

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

Properties

  • Form

    Liquid
  • Storage instructions

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

    pH: 7.2
    Constituent: PBS
  • Carrier free

    Yes
  • Concentration information loading...
  • Purity

    Protein A purified
  • Clonality

    Monoclonal
  • Clone number

    EPR20535
  • Isotype

    IgG
  • Research areas

    • Neuroscience
    • Neurology process
    • Neurodegenerative disease
    • Parkinson's disease
    • Synuclein
    • Neuroscience
    • Neurology process
    • Neurodegenerative disease
    • Alzheimer's disease
    • Tangles & Tau
    • Neuroscience
    • Diseases
    • Neuroscience
    • Processes

Associated products

  • Alternative Versions

    • Anti-Alpha-synuclein antibody [EPR20535] (ab212184)
  • Compatible Secondaries

    • Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab205718)
    • Goat Anti-Rabbit IgG H&L (HRP) (ab97051)
  • Conjugation kits

    • FITC Conjugation Kit (Fast) - Lightning-Link® (ab188285)
  • Isotype control

    • Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
  • Recombinant Protein

    • Recombinant Human Alpha-synuclein protein aggregate (Active) (ab218819)
  • Related Products

    • VeriBlot for IP Detection Reagent (HRP) (ab131366)
    • Antigen Retrieval Buffer (100X Tris-EDTA Buffer, pH 9.0) (ab93684)

Applications

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab225866 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 at an assay dependent concentration. Detects a band of approximately 18 kDa (predicted molecular weight: 14 kDa).
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
IHC-Fr
Use at an assay dependent concentration.

Antigen retrieval: Heated citrate solution (10mM citrate pH 6.0 + 0.05% Tween-20).

IP
Use at an assay dependent concentration.
Notes
WB
Use at an assay dependent concentration. Detects a band of approximately 18 kDa (predicted molecular weight: 14 kDa).
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
IHC-Fr
Use at an assay dependent concentration.

Antigen retrieval: Heated citrate solution (10mM citrate pH 6.0 + 0.05% Tween-20).

IP
Use at an assay dependent concentration.

Target

  • Function

    May be involved in the regulation of dopamine release and transport. Induces fibrillization of microtubule-associated protein tau. Reduces neuronal responsiveness to various apoptotic stimuli, leading to a decreased caspase-3 activation.
  • Tissue specificity

    Expressed principally in brain but is also expressed in low concentrations in all tissues examined except in liver. Concentrated in presynaptic nerve terminals.
  • Involvement in disease

    Genetic alterations of SNCA resulting in aberrant polymerization into fibrils, are associated with several neurodegenerative diseases (synucleinopathies). SNCA fibrillar aggregates represent the major non A-beta component of Alzheimer disease amyloid plaque, and a major component of Lewy body inclusions. They are also found within Lewy body (LB)-like intraneuronal inclusions, glial inclusions and axonal spheroids in neurodegeneration with brain iron accumulation type 1.
    Parkinson disease 1
    Parkinson disease 4
    Dementia Lewy body
  • Sequence similarities

    Belongs to the synuclein family.
  • Domain

    The 'non A-beta component of Alzheimer disease amyloid plaque' domain (NAC domain) is involved in fibrils formation. The middle hydrophobic region forms the core of the filaments. The C-terminus may regulate aggregation and determine the diameter of the filaments.
  • Post-translational
    modifications

    Phosphorylated, predominantly on serine residues. Phosphorylation by CK1 appears to occur on residues distinct from the residue phosphorylated by other kinases. Phosphorylation of Ser-129 is selective and extensive in synucleinopathy lesions. In vitro, phosphorylation at Ser-129 promoted insoluble fibril formation. Phosphorylated on Tyr-125 by a PTK2B-dependent pathway upon osmotic stress.
    Hallmark lesions of neurodegenerative synucleinopathies contain alpha-synuclein that is modified by nitration of tyrosine residues and possibly by dityrosine cross-linking to generated stable oligomers.
    Ubiquitinated. The predominant conjugate is the diubiquitinated form.
    Acetylation at Met-1 seems to be important for proper folding and native oligomeric structure.
  • Cellular localization

    Cytoplasm, cytosol. Membrane. Nucleus. Cell junction, synapse. Secreted. Membrane-bound in dopaminergic neurons.
  • Target information above from: UniProt accession P37840 The UniProt Consortium
    The Universal Protein Resource (UniProt) in 2010
    Nucleic Acids Res. 38:D142-D148 (2010) .

    Information by UniProt
  • Database links

    • Entrez Gene: 6622 Human
    • Entrez Gene: 20617 Mouse
    • Entrez Gene: 29219 Rat
    • Omim: 163890 Human
    • SwissProt: P37840 Human
    • SwissProt: O55042 Mouse
    • SwissProt: P37377 Rat
    • Unigene: 21374 Human
    • Unigene: 17484 Mouse
    • Unigene: 1827 Rat
    see all
  • Alternative names

    • Alpha synuclein antibody
    • Alpha-synuclein antibody
    • Alpha-synuclein, isoform NACP140 antibody
    • alphaSYN antibody
    • MGC105443 antibody
    • MGC110988 antibody
    • MGC127560 antibody
    • MGC64356 antibody
    • NACP antibody
    • Non A beta component of AD amyloid antibody
    • Non A4 component of amyloid antibody
    • Non A4 component of amyloid precursor antibody
    • Non-A beta component of AD amyloid antibody
    • Non-A-beta component of alzheimers disease amyloid , precursor of antibody
    • Non-A4 component of amyloid precursor antibody
    • Non-A4 component of amyloid, precursor of antibody
    • OTTHUMP00000218549 antibody
    • OTTHUMP00000218551 antibody
    • OTTHUMP00000218552 antibody
    • OTTHUMP00000218553 antibody
    • OTTHUMP00000218554 antibody
    • PARK 1 antibody
    • PARK 4 antibody
    • PARK1 antibody
    • PARK4 antibody
    • Parkinson disease (autosomal dominant, Lewy body) 4 antibody
    • Parkinson disease familial 1 antibody
    • SNCA antibody
    • Snca synuclein antibody
    • Snca synuclein, alpha (non A4 component of amyloid precursor) antibody
    • SYN antibody
    • Synuclein alpha antibody
    • Synuclein alpha 140 antibody
    • Synuclein, alpha (non A4 component of amyloid precursor) antibody
    • SYUA_HUMAN antibody
    see all

Images

  • Western blot - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Western blot - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    All lanes : Anti-Alpha-synuclein antibody [EPR20535] (ab212184) at 1/1000 dilution

    Lane 1 : Human Alpha-synuclein recombinant protein
    Lane 2 : Human Beta-synuclein recombinant protein

    Lysates/proteins at 0.01 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 14 kDa
    Observed band size: 18 kDa why is the actual band size different from the predicted?


    Exposure time: 8 seconds


    This data was developed using ab212184, the same antibody clone in a different buffer formulation

    Blocking/Dilution buffer: 5% NFDM/TBST.

    Human Alpha-synuclein recombinant protein contain aa1-140 with His-tag. Human Beta-synuclein recombinant protein contain aa1-134 with His-tag.

  • Western blot - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Western blot - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    All lanes : Anti-Alpha-synuclein antibody [EPR20535] (ab212184) at 1/1000 dilution

    Lane 1 : Human cerebellum lysate
    Lane 2 : Human brain lysate
    Lane 3 : Mouse brain lysate
    Lane 4 : Rat brain lysate

    Lysates/proteins at 20 µg per lane.

    Secondary
    All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

    Predicted band size: 14 kDa
    Observed band size: 18 kDa why is the actual band size different from the predicted?


    Exposure time: 5 seconds


    This data was developed using ab212184, the same antibody clone in a different buffer formulation

    Blocking/Dilution buffer: 5% NFDM/TBST.

    The observed molecular weight is consistent with the literature (PMID: 11739566).

  • Western blot - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Western blot - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    All lanes : Anti-Alpha-synuclein antibody [EPR20535] (ab212184) at 1/1000 dilution

    Lane 1 : Wild-type U-87 MG cell lysate
    Lane 2 : SNCA knockout U-87 MG cell lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 14 kDa
    Observed band size: 16 kDa why is the actual band size different from the predicted?



    This data was developed using ab212184, the same antibody clone in a different buffer formulation:

    False colour image of Western blot: Anti-Alpha-synuclein antibody [EPR20535] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab212184 was shown to bind specifically to Alpha-synuclein. A band was observed at 16 kDa in wild-type U-87 MG cell lysates with no signal observed at this size in SNCA knockout cell line ab282333 (knockout cell lysate ab283006). To generate this image, wild-type and SNCA knockout U-87 MG cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.

  • Western blot - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Western blot - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)

    This data was developed using ab212184, the same antibody clone in a different buffer formulation:

    Lane 1: Wild type HAP1 whole cell lysate (20 µg)
    Lane 2: SNCA (alpha Synuclein) knockout HAP1 whole cell lysate (20 µg)
    Lane 3: Human brain whole tissue lysate (20 µg)
    Lane 4: SH-SY5Y whole cell lysate (20 µg)

    Lanes 1 - 4: Merged signal (red and green). Green - ab212184 observed at 14 kDa. Red - loading control, ab9484, observed at 37 kDa.

    ab212184 was shown to specifically react with SNCA (alpha Synuclein) in wild type cells as signal was lost in SNCA (alpha Synuclein) knockout cells. Wild-type and SNCA (alpha Synuclein) knockout samples were subjected to SDS-PAGE. ab212184 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at a 1/1000 dilution and 1/20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Immunohistochemical analysis of paraffin-embedded human glioma tissue labeling Alpha-synuclein with ab212184 at 1/16000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
     
    Cytoplasmic staining on human glioma [PMID: 22112368].
    Counter stained with Hematoxylin.
     
    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab212184).

    Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Immunohistochemical analysis of paraffin-embedded mouse cerebral cortex tissue labeling Alpha-synuclein with ab212184 at 1/16000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
     
    Cytoplasmic staining on mouse cerebral cortex [PMID: 22112368].

    Counter stained with Hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab212184).

    Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Immunohistochemical analysis of paraffin-embedded rat cerebral cortex tissue labeling Alpha-synuclein with ab212184 at 1/16000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
     
    Cytoplasmic staining on rat cerebral cortex [PMID: 22112368].

    Counter stained with Hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab212184).

    Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Immunohistochemical analysis of paraffin-embedded human kidney tissue labeling Alpha-synuclein with ab212184 at 1/16000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
     
    Negative control: No staining on human kidney. [PMID: 14997013].

    Counter stained with Hematoxylin.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab212184).

    Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

  • Immunohistochemistry (Frozen sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Immunohistochemistry (Frozen sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)

    Immunohistochemical analysis of 4% paraformaldehyde-fixed, 0.2% Triton X-100 permeabilized frozen mouse hippocampus tissue labeling Alpha-synuclein with ab212184 at 1/100 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

    Cytoplasmic staining on mouse hippocampus (PMID: 22112368).

    The nuclear counterstain is DAPI (blue).

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) at 1/1000 dilution.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab212184).

  • Immunoprecipitation - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Immunoprecipitation - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)

    Alpha-synuclein was immunoprecipitated from 0.35 mg of mouse brain lysate with ab212184 at 1/30 dilution.

    Western blot was performed from the immunoprecipitate using ab212184 at 1/1000 dilution.

    VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution.

    Lane 1: Mouse brainlysate 10ug (Input).

    Lane 2: ab212184 IP in mouse brain lysate.

    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab212184 in mouse brain lysate.

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

    Exposure time: 1 second.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab212184).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Immunohistochemical analysis of paraffin-embedded human cerebral cortex tissue labeling Alpha-synuclein with ab212184 at 1/16000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.
     
    Cytoplasmic staining on human cerebral cortex [PMID: 22112368].

    Counter stained with Hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab212184).

     

    Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

  • Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)
    Anti-Alpha-synuclein antibody [EPR20535] - BSA and Azide free (ab225866)

Protocols

To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.

Click here to view the general protocols

Datasheets and documents

  • Datasheet download

    Download

Certificate of Compliance

To download a Certificate of Compliance, please enter your Lot number below:

References (0)

Publishing research using ab225866? Please let us know so that we can cite the reference in this datasheet.

ab225866 has not yet been referenced specifically in any publications.

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