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Enhanced validation of antibodies for research applications

Author

Listed:
  • Fredrik Edfors

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

  • Andreas Hober

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

  • Klas Linderbäck

    (KTH - Royal Institute of Technology)

  • Gianluca Maddalo

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

  • Alireza Azimi

    (Karolinska University Hospital)

  • Åsa Sivertsson

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

  • Hanna Tegel

    (KTH - Royal Institute of Technology)

  • Sophia Hober

    (KTH - Royal Institute of Technology)

  • Cristina Al-Khalili Szigyarto

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

  • Linn Fagerberg

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

  • Kalle Feilitzen

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

  • Per Oksvold

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

  • Cecilia Lindskog

    (Uppsala University)

  • Björn Forsström

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology)

  • Mathias Uhlen

    (KTH - Royal Institute of Technology
    KTH - Royal Institute of Technology
    Technical University of Denmark)

Abstract

There is a need for standardized validation methods for antibody specificity and selectivity. Recently, five alternative validation pillars were proposed to explore the specificity of research antibodies using methods with no need for prior knowledge about the protein target. Here, we show that these principles can be used in a streamlined manner for enhanced validation of research antibodies in Western blot applications. More than 6,000 antibodies were validated with at least one of these strategies involving orthogonal methods, genetic knockdown, recombinant expression, independent antibodies, and capture mass spectrometry analysis. The results show a path forward for efforts to validate antibodies in an application-specific manner suitable for both providers and users.

Suggested Citation

  • Fredrik Edfors & Andreas Hober & Klas Linderbäck & Gianluca Maddalo & Alireza Azimi & Åsa Sivertsson & Hanna Tegel & Sophia Hober & Cristina Al-Khalili Szigyarto & Linn Fagerberg & Kalle Feilitzen & P, 2018. "Enhanced validation of antibodies for research applications," Nature Communications, Nature, vol. 9(1), pages 1-10, December.
    • Handle: RePEc:nat:natcom:v:9:y:2018:i:1:d:10.1038_s41467-018-06642-y
    DOI: 10.1038/s41467-018-06642-y
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    Cited by:

    1. Shideh Mirhadi & Shirley Tam & Quan Li & Nadeem Moghal & Nhu-An Pham & Jiefei Tong & Brian J. Golbourn & Jonathan R. Krieger & Paul Taylor & Ming Li & Jessica Weiss & Sebastiao N. Martins-Filho & Vibh, 2022. "Integrative analysis of non-small cell lung cancer patient-derived xenografts identifies distinct proteotypes associated with patient outcomes," Nature Communications, Nature, vol. 13(1), pages 1-17, December.
    2. Joshua D. Spitzberg & Scott Ferguson & Katherine S. Yang & Hannah M. Peterson & Jonathan C. T. Carlson & Ralph Weissleder, 2023. "Multiplexed analysis of EV reveals specific biomarker composition with diagnostic impact," Nature Communications, Nature, vol. 14(1), pages 1-12, December.
    3. Fei Wang & Peiwen Ding & Xue Liang & Xiangning Ding & Camilla Blunk Brandt & Evelina Sjöstedt & Jiacheng Zhu & Saga Bolund & Lijing Zhang & Laura P. M. H. Rooij & Lihua Luo & Yanan Wei & Wandong Zhao , 2022. "Endothelial cell heterogeneity and microglia regulons revealed by a pig cell landscape at single-cell level," Nature Communications, Nature, vol. 13(1), pages 1-18, December.

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