Tuesday, February 1, 2011

0 Paper On ELISA And Immunoblot Differences

The Lyme patient community has been saying all along that blood tests for Lyme Disease are inaccurate and vary in specificity and sensitivity for some time.

This recent paper is more data in support of this complaint.

Full text version: http://www.springerlink.com/content/w64170894v08654g/fulltext.html

Eur J Clin Microbiol Infect Dis. 2011 Jan 27; [Epub ahead of print]

C. W. Ang1 Contact Information, D. W. Notermans2, M. Hommes1, A. M. Simoons-Smit1 and T. Herremans2
(1) VUMC, Amsterdam, The Netherlands
(2) Centre for Infectious Disease Control Netherlands, National Institute for Public Health and the Environment (RIVM), Bilthoven, The Netherlands
Received: 21 July 2010 Accepted: 1 January 2011 Published online: 27 January 2011

Large differences between test strategies for the detection of anti-Borrelia antibodies are revealed by comparing eight ELISAs and five immunoblots.

ABSTRACT

We investigated the influence of assay choice on the results in a two-tier testing algorithm for the detection of anti-Borrelia antibodies.

Eighty-nine serum samples from clinically well-defined patients were tested in eight different enzyme-linked immunosorbent assay (ELISA) systems based on whole-cell antigens, whole-cell antigens supplemented with VlsE and assays using exclusively recombinant proteins.

A subset of samples was tested in five immunoblots: one whole-cell blot, one whole-cell blot supplemented with VlsE and three recombinant blots.

The number of IgM- and/or IgG-positive ELISA results in the group of patients suspected of Borrelia infection ranged from 34 to 59%.

The percentage of positives in cross-reactivity controls ranged from 0 to 38%.

Comparison of immunoblots yielded large differences in inter-test agreement and showed, at best, a moderate agreement between tests.

Remarkably, some immunoblots gave positive results in samples that had been tested negative by all eight ELISAs.

The percentage of positive blots following a positive ELISA result depended heavily on the choice of ELISA-immunoblot combination.

We conclude that the assays used to detect anti-Borrelia antibodies have widely divergent sensitivity and specificity.

The choice of ELISA-immunoblot combination severely influences the number of positive results, making the exchange of test results between laboratories with different methodologies hazardous.



It would be good to see an analysis like this on all North American ELISA and immunoblot tests - this paper focuses on European tests. 


The full text for this paper showed that not only does two-tier testing miss positive cases, but that  specificity between immunoblots varies greatly and can miss positive cases. The implication is that using some immunoblot tests to confirm positive ELISA tests may be a flawed approach due to differences between immunoblots. 


The discussion section states that, "Theoretically, the use of recombinant antigens should lead to increased specificity and, possibly, increased sensitivity as well. This does not seem to be true for the currently available ELISAs and immunoblots for the detection of anti-Borrelia antibodies. We could not find a clear relationship between the fraction of positive tests, the specificity and the nature of the antigen used for the serological tests. ELISAs using sonicated whole-cell antigens can be sensitive and specific, while recombinant ELISAs may lack specificity. Therefore, manufacturer claims for the superior performance of assays using recombinant antigens for the detection of Borrelia antibodies must be interpreted with caution."

Problems in detection occur with both the first tier and second tier of testing, and results are further  affected by how early in the infection the patient is tested and whether or not the patient has had any antibiotics recently.


This seems to confirm the need for more awareness of how sensitive and specific each lab's tests are at each tier, and to rely more on clinical diagnosis and patient history to determine treatment.

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