{"id":2323,"date":"2017-08-15T17:35:00","date_gmt":"2017-08-15T21:35:00","guid":{"rendered":"https:\/\/www.med.unc.edu\/flowcytometry\/resources-2\/antibody-validation-and-reproducibility-resources-1\/"},"modified":"2019-03-19T09:56:07","modified_gmt":"2019-03-19T13:56:07","slug":"antibody-validation-and-reproducibility-resources","status":"publish","type":"page","link":"https:\/\/www.med.unc.edu\/flowcytometry\/resources\/antibody-validation-and-reproducibility-resources\/","title":{"rendered":"Antibody Validation and Reproducibility resources"},"content":{"rendered":"
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First<\/strong> – be sure to search by lot number<\/strong> in addition to catalog number!!\u00a0 Just because one lot was validated, it does not validate another lot!!<\/h5>\n<\/blockquote>\n

Antibody Registry\u00a0 \u00a0<\/strong><\/p>\n

The Antibody Registry includes commercial antibodies from hundreds of commercial vendors and thousands of individual labs. However, many of the links don’t go anywhere useful. Give it a try:\u00a0 \u00a0http:\/\/antibodyregistry.org\/<\/a><\/h5>\n

Or use BenchSci<\/strong><\/span>\u00a0\u00a0https:\/\/landing.benchsci.com\/academic<\/a>\u00a0 \u00a0 ‘We use AI to screen the literature for published antibodies so you don\u2019t have to.’<\/p>\n

<\/h4>\n

Companies:\u00a0<\/strong> practices vary immensely – do your homework before buying!!<\/h4>\n

BD Biosciences: [awaiting response<\/em>]<\/strong><\/p>\n

BioLegend<\/strong> page on Antibody Validation Challenges: Reproducibility and Validation Webpage, including steps to improve reproducibility:\u00a0\u00a0https:\/\/www.biolegend.com\/reproducibility<\/a><\/p>\n<\/div>\n

Capricobio:\u00a0\u00a0<\/strong>\u2018We are an ISO certified cGMP facility and validate every lot of antibody that is produced. We validate our antibodies through cell binding specificity COA and other validation information can be obtained at customer request for their lot of interest.\u2019<\/p>\n

Cell Signaling Technology<\/strong> Antibody Validation:\u00a0\u00a0https:\/\/www.cellsignal.com\/contents\/our-approach\/cst-antibody-validation-principles\/ourapproach-validation-principles<\/a><\/p>\n

Jackson Immuno Research\u00a0 (mostly secondary antibodies)<\/strong>:\u00a0 ‘Most of JIR\u2019s products are polyclonal secondary antibodies, and because each preparation can recognize many epitopes on the primary antibody targets, the secondaries will react with the primaries wherever they are bound. If the primary has bound its target antigen (under any fixation condition), the secondary antibody will recognize it.\u00a0Our Quality Assurance testing is designed to confirm that the antibodies behave as we describe them. The ability of the secondary antibody to bind to its intended target is confirmed by immunoelectrophoresis and\/or a solid-phase immunoaffinity assay (target protein coupled to agarose beads). Each antibody purification event is also tested by a sensitive ELISA assay to confirm that products we describe as having \u201cminimal cross-reactivity\u201d to other species meet our standard (<1% reactivity as compared with the intended target). We have similar standards for products that are Fc or F(ab\u2019)2 fragment specific, and for our anti-mouse subclass-specific reagents. All the products are tested in-process, and retested after vialing \/ lyophilization to confirm their performance.’\u00a0\u00a0https:\/\/www.jacksonimmuno.com\/<\/a><\/p>\n

Milllipore\/Sigma-Aldrich\u00a0\u00a0<\/strong>Antibody Enhanced Validation \u2013 Additional Validation Data to Help Ensure Specificity and Performance<\/a><\/p>\n

MiltenyiBiotec<\/strong> – Miltenyi Biotec has engineered new recombinant antibodies designed specifically for flow cytometry: REAfinity\u2122 Recombinant Antibodies. These antibodies eliminate binding to Fc receptors and their recombinant production improves lot-to-lot consistency and reproducibility across multiple experiments. All antibodies are validated to work in flow cytometry. Additionally, enhanced validation data for antibody-specificity, sensitivity, and reproducibility are available on our website under each specific antibody clone. Please visit www.miltenyibiotec.com\/reafinity<\/a> for more information.<\/p>\n

Santa Cruz<\/strong>\u00a0 ‘We have validated all of our antibodies but only in Western Blotting. We have not done any validations for Immunofluorescence, Immuno Histochemistry, ELISA, or any other applications.’\u00a0 \u00a0– DO NOT USE SANTA CRUZ ANTIBODIES FOR FLOW CYTOMETRY UNTIL YOU HAVE VALIDATED THEM!!<\/span><\/p>\n

ThermoFisher<\/strong><\/p>\n

https:\/\/www.thermofisher.com\/us\/en\/home\/life-science\/antibodies\/invitrogen-antibody-validation.html<\/a><\/p>\n

 <\/p>\n

UNDER CONSTRUCTION:<\/strong><\/h5>\n

AbCam, BD<\/p>\n

Other tips and resources:\u00a0<\/strong><\/h4>\n

Information from the experts on antibody validation from BenchSci\u00a0\u00a0https:\/\/blog.benchsci.com\/reproducible-research<\/a><\/p>\n

All antibodies should be validated prior to reporting of results.\u00a0Recommendations for research using antibodies<\/a>\u00a0(page 7-8) \u2018Although vendor-supplied technical information may help investigators select reagents such as antibodies, this information is insufficient for validation\u2019.<\/h5>\n
Antibody Validation: Standards, Policies and Practices\u00a0Workshop Report<\/a>\u00a02016<\/h5>\n
Best practices for using antibodies for flow cytometry:<\/strong><\/h5>\n
    \n
  1. \n
    Always Titrate<\/h5>\n<\/li>\n
  2. \n
    Validate Specificity (If manufacturer has not done this, then you need to!!)<\/h5>\n<\/li>\n
  3. \n
    Integrate critical controls like Fluorescence Minus One controls (FMO)<\/h5>\n<\/li>\n
  4. \n
    See: Uhlen et al.,\u00a0A Proposal for Validation of Antibodies<\/a>, Nature Methods (2016)<\/h5>\n<\/li>\n
  5. \n
    More at:\u00a0https:\/\/expertcytometry.com\/4-steps-to-validate-flow-cytometry-antibodies-and-improve-reproducibility\/<\/a><\/h5>\n<\/li>\n
  6. \n
    Also on the EuroMab network:<\/h5>\n
    Guidelines Website:\u00a0https:\/\/www.euromabnet.com\/guidelines\/index.php<\/a>\u00a0\u00a0 Guidelines Manuscript:\u00a0https:\/\/www.ncbi.nlm.nih.gov\/pubmed\/26418356<\/a>\u00a0\u00a0 Pos\/Neg Controls Website:\u00a0https:\/\/www.euromabnet.com\/guidelines\/positive-negative-controls.php<\/a>\u00a0\u00a0 Additional Articles about Ab Validation:\u00a0https:\/\/www.euromabnet.com\/guidelines\/articles-about-antibody-validation.php<\/a><\/h5>\n<\/li>\n<\/ol>\n
    <\/h5>\n
    <\/h5>\n

    Attn: other antibody manufacturers who wish to be added to this list, please provide links your antibody validation process and where investigators can access the validation data for each lot. I will be happy to include you in this list!\u00a0 ncfisher@med.unc.edu.<\/p>\n

    Investigators: if a company you work with is not fully linked on this list, be sure to research your antibody source and validation before<\/em> purchase!!<\/p>\n

     <\/p>\n","protected":false},"excerpt":{"rendered":"

    First – be sure to search by lot number in addition to catalog number!!\u00a0 Just because one lot was validated, it does not validate another lot!! Antibody Registry\u00a0 \u00a0 The Antibody Registry includes commercial antibodies from hundreds of commercial vendors and thousands of individual labs. However, many of the links don’t go anywhere useful. Give … Read more<\/a><\/p>\n","protected":false},"author":20161,"featured_media":0,"parent":2263,"menu_order":8,"comment_status":"closed","ping_status":"closed","template":"","meta":{"_acf_changed":false,"layout":"","cellInformation":"","apiCallInformation":"","footnotes":"","_links_to":"","_links_to_target":""},"class_list":["post-2323","page","type-page","status-publish","hentry","odd"],"acf":[],"_links_to":[],"_links_to_target":[],"_links":{"self":[{"href":"https:\/\/www.med.unc.edu\/flowcytometry\/wp-json\/wp\/v2\/pages\/2323","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/www.med.unc.edu\/flowcytometry\/wp-json\/wp\/v2\/pages"}],"about":[{"href":"https:\/\/www.med.unc.edu\/flowcytometry\/wp-json\/wp\/v2\/types\/page"}],"author":[{"embeddable":true,"href":"https:\/\/www.med.unc.edu\/flowcytometry\/wp-json\/wp\/v2\/users\/20161"}],"replies":[{"embeddable":true,"href":"https:\/\/www.med.unc.edu\/flowcytometry\/wp-json\/wp\/v2\/comments?post=2323"}],"version-history":[{"count":0,"href":"https:\/\/www.med.unc.edu\/flowcytometry\/wp-json\/wp\/v2\/pages\/2323\/revisions"}],"up":[{"embeddable":true,"href":"https:\/\/www.med.unc.edu\/flowcytometry\/wp-json\/wp\/v2\/pages\/2263"}],"wp:attachment":[{"href":"https:\/\/www.med.unc.edu\/flowcytometry\/wp-json\/wp\/v2\/media?parent=2323"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}