HCP Coverage Analysis

  • Evaluation and comparison of ELISA kits
  • Low amount of antibody required - 1 mg
  • Alternative to 2D Gels & Western Blotting

Do you know how well your ELISA covers the HCPs in your sample?

If you need to evaluate and qualify the antibodies of a generic or process specific HCP ELISA we can help you. We use an advanced MS-based coverage analysis method (ELISA-MS) which mimics the conditions of the ELISA, for more reliable and accurate results:

In this analysis, the host cell proteins in a mock lysate or an early process sample are immunocaptured u...

Do you know how well your ELISA covers the HCPs in your sample?

If you need to evaluate and qualify the antibodies of a generic or process specific HCP ELISA we can help you. We use an advanced MS-based coverage analysis method (ELISA-MS) which mimics the conditions of the ELISA, for more reliable and accurate results:

In this analysis, the host cell proteins in a mock lysate or an early process sample are immunocaptured using the ELISA antibodies and then identified by LC-MS/MS. As a result, you get not only the total sum of the residual HCPs, but also a list of the HCPs recognized by the ELISA antibodies.

How does MS analysis compare to 2D PAGE and Western blot?
ELISA-MS is a proven method recognized as a better alternative to spot counting in 2D gel electrophoresis and Western blot, because it provides much more detailed and accurate information. An Immunocapture LC-MS analysis (ELISA-MS) will thus expand your knowledge of the HCP-ELISA’s specificity and applicability to your biopharmaceutical. It may also reduce the risk of project delays due to poorly documented HCP ELISA.

A standard ELISA-MS HCP coverage analysis includes

  • ELISA setup for optimal antibody-HCP binding.
  • Immunocapture with ELISA polyclonal antibodies.
  • Identification of both total HCPs and affinity purified HCPs by sensitive LC-MS/MS.
  • Reporting of HCPs covered by antibodies and also a list of total HCPs in the mock lysate or early proces sample.

Send us a description of your project and we will be glad to discuss it with you.

MoreLess details

The process:

  • 1 Contact us to discuss your project and receive a project proposal
  • 2 Analysis phase lead by Alphalyse appointed principal investigator
  • 3 Report w. list of recognized HCPs and total HCP coverage
  • 4 Follow up by phone or email

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We offer customized solutions, contact us to discuss your project.

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Testimonials

  • "They handled the project professionally and rapidly, and the report that was very well written, clearly explaining the findings."

    Targovax ASA, Kristiina Hyvärinen – Director, QC, viral products

  • "Using the Alphalyse LC-MS/MS coverage method in HCP ELISA selection, we estimate a savings of approximately $1M and, likely, one year of development time"

    SAVARA Aps, Horsholm, Denmark Lars Skriver, Senior Science Officer

Knowledge center

More information

We would like to help you as much as possible with your project and therefore provide several kinds of customer support:

Technical details

The HCP coverage analysis (ELISA-MS) includes:

  1. First, ELISA setup for optimal antibody-HCP binding.
  2. Next, capture of HCPs (antigens) to ELISA antibodies immobilized in 96-well plates.
  3. Finally, digest and LC-MS analysis for HCP identification.
ELISA HCP coverage analysis illustration

Illustration of the ELISA-MS method

An aliquot of the antigen sample (a mock/null cell lysate or early process sample) is digested under similar conditions in parallel without antibodies and serves as a measurement of total protein content to identify all HCPs. This is done to compare the number of HCPs bound to ELISA antibodies with the number of HCPs in the original sample (total protein sample).

Applications of ELISA-MS

We have performed a wide range of projects evaluating commercial/generic, process-specific and platform assays from common HCP kit manufacturers like Biogenes, Canopy, Cygnus, Creative Diagnostics, Rockland, etc.

The hcp coverage analysis can evaluate all HCP ELISAs, e.g. those based on the common expression systems:

  • Yeast: S. cerevisiae, P. pastoris
  • Bacterial: E. coliL. lactisP. fluorescens, S. aureus
  • Insect: S. frugiperda (SF9)
  • Mammalian: HEK293, CHO, HeLa, MRC5, GS-NS0, Vero cell, SP2/0, BHK, A549, MDCK

Along with specific process residuals, such as human serum albumin (HSA), bovine serum albumin (BSA) and Protein A.

Sample preparation

Sample requirements for MS-ELISA

HCP coverage analysis using mass spectrometry technology requires both ELISA antibodies and a mock cell lysate or early process sample.

  1. Antibodies

    1. The ELISA optimization and immunocapture analysis requires typically a minimum of 1 mg of antibodies
    2. The samples should not contain large amounts of detergents
    3. The concentration should be >0.5 µg/µl
    4. Use a Eppendorf Safe Lock tube or similar tubes
    5. Freeze or refrigerate to +4 oC for cold shipment of the liquid sample
  2. MOCK/early process sample

    1. The mock/early process sample requires typically a minimum of 100 µg of protein material
    2. The samples should not contain large amounts of detergents
    3. The concentration should be >0.5 µg/µl
    4. Use a Eppendorf Safe Lock tube supplied, or similar tubes from your lab
    5. Freeze or refrigerate to +4 oC for cold shipment of the liquid sample

Please contact Alphalyse for sample requirements before shipment.

Meet the experts

For this type of analysis our experts include:

Do you need help?
Solveig Beck Nielsen

PhD in Molecular Biology

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Do you need help?
Katrine Pilely

PhD in Biomedicine and BSc in Biology

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Do you need help?
Flemming Holm Bergholt

Biomedical Laboratory Scientist

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