Talk:ELISA

Fluorogenic and Electrochemiluminescent Substrates reference

The reference: Toedter G, Hayden K, Wagner C, Brodmerkel C (January 2008). "Simultaneous Detection of Eight Analytes in Human Serum by Two Commercially Available Platforms for Multiplex Cytokine Analysis". Clinical and Vaccine Immunology 15: 42-48. PMID 18003817. does not support the statement: "though newer assays employ fluorogenic and electrochemiluminescent substrates enabling much higher sensitivity" Rather the paper is reporting that there appears to be significant inter-company differences in results for a given cytokine, using cytokine multiplex ELISA. —Preceding unsigned comment added by Gregnz (talk • contribs) 07:15, 26 January 2010 (UTC)

Hey Gregnz, You were right about the reference. I was playing it a little fast and loose when I initially wrote this section. I did a quick rewrite, and I think both the language and the referencing is better. I should probably note that I claim some expertese in this field (this isn't to say I don't want to be challenged, but just that I'm not a laymen reguritating from references).

Let me know if you feel this rewrite addresses your concerns. NickCT (talk) 14:51, 26 January 2010 (UTC)

Hi NickCT, thanks. I've used fluorogenic ELISA (which are enzyme-linked, and have been around since the 80s), and I was aware of multiplexes, but the RT-PCR and electrochemiluminescent 'ELISA' are news to me, as was the improved sensitivity. I would like to link to a peer-reviewed report on the improved sensitivity of these methods, and the current reference is published by the company that makes the allegedly most sensitive assay. Of course, neither of these reasons mean that the claims aren't true. I've done a quick google, and I think this: "Sensitivity by combination: immuno-PCR and related technologies. Michael Adler, RonWacker and Christof M. Niemeyer DOI: 10.1039/b718587c Analyst, 2008, 133, 702–718" is a nice review on immuno-PCR that also deals with the improved sensitivity. But I don't know how to add references!

Do you think we could improve on the ELISA article in general, including sections on the newer techniques?

Gregnz (talk) 20:39, 27 January 2010 (UTC)

I agree this article could use a significant rewrite. I'm not sure I have the time though. I'll keep it in mind though. NickCT (talk) 14:06, 28 January 2010 (UTC)

SPAM

Hi, I was just researching some information and came across someone who edited the application part of this article with complete nonsense about ELISA being a "super cute" virus that was attacking your brain. I undid the change. -Asrrin29 —Preceding unsigned comment added by 71.13.85.116 (talk) 12:03, 26 June 2008 (UTC)

ELISA

ELISA and EIA are closley related, but different things - ELISA is a swedish invention and EIA is a Dutch invention

The article is completely impenetrable to laypersons. Not only are the explanations immersed in jargon, but there is no plainly stated purpose of ELISA. Why would one use it? What for? Are there newer techniques that (may) make it obsolete?

Does anyone have information on "false positive" and "false negative" rates and interpretations? What are these rates? What situations can lead to a false positive or a false negative ... both test errors as well as clinical causes.

I suspect that false postives and negatives are related to the specific test being run by ELISA, rather than the general technique. I think that that individual reagents and their conditions are where problems would mostly arise from. Dancarney 22:44, 11 July 2006 (UTC)

You are correct. Not only that, but false positive and false negative rates are also dependent upon who manufactured th kit, what batch the kit came from, and a large number of other factors. Considering that the antibodies used in ELISA usually come from injecting rabbits or goats and collecting the antibodies, quality control c

The rate of false positives and negatives relate to the application of an ELISA result to the discrimination between categories (e.g. presence or absence of disease/drugs of abuse), rather than anything intrinsic to the ELISA system. The rate of false positives and negatives(along with sensitivity and specificity) will vary between applications by the variance of the diagnostic test, but also the cut off level selected, among other things. The accuracy of the results will depend on the things you mention above. The coefficient of variance (%CV) is a measure of difference between samples tested in duplicate, and is one of the ways that the accuracy of an ELISA is checked. Another way is seeing how accurately an ELISA result matches up when testing a sample with a known amount what ever you are looking for. Gregnz (talk) 07:11, 26 January 2010 (UTC)

ELISA-sandwich.svg

This picture is confusing and do not correspond to the written description of a sandwich ELISA. It looks like a mixture between a indirect and sandwich ELISA. I think the enzyme-linked antibody binds to the antigen directly. No "detecting antibody" is needed, so the third step described in the picture, "(3) detecting antibody is added, and binds to antigen;" is wrong and should be deleted. /Izor 130.243.248.239 17:08, 15 October 2006 (UTC)

I agree that the text and the image do not match up. I think the image actually represents a very common methodology used for ELISA (at least in my experience). You're right that a detecting antibody may not be needed if the secondary antibody is enzyme-conjugated, but in practice the conjugation is expensive and thus can be much more practically done by using an extra step with a detecting antibody. Anyway, I've been meaning to fix up and clarify the text for a while, but I haven't had time to do it. Please go ahead if you have the time, though...Be bold!  — JVinocur (talk • contribs) 17:30, 16 October 2006 (UTC)

I've attempted to make this clearer. If someone would like to rewrite so that it reads more nicely, they could then delete these three posts. --Username132 (talk) 18:15, 24 October 2006 (UTC)

I've printed off this article to refer to with a university project I'm doing, however the image prints off with a black background, and all that shows is the blue and red shapes. I've tried saving the image and changing it to jpeg or similar, but same happens? Is there any reason for this? can anyone help?! Thanks Alex 17:12, 24 November 2006 (UTC)

Not sure what to tell you, unfortunately...you might try using a different web browser, I guess?  — JVinocur (talk • contribs) 23:44, 26 November 2006 (UTC)

It's OK now, I used the ingenious, and extremely technical technique of pressing 'print screen' to get a screen shot of the image! Thanks for your help anyway, it probably is my browser; I use firefox. Alex 23:29, 10 December 2006 (UTC)

precautions while doing ELISA

tell me the precautions

Response to Precautions: What exactly are you looking for in terms of precautionary steps? Safety? Potential sources of error? That's a tad vague there. —Preceding unsigned comment added by 69.123.232.139 (talk) 15:34, 8 September 2007 (UTC)

RIA

I changed this:

Prior to the development of the EIA/ELISA, immunoassays were conducted using radioactively-labeled antigens or antibodies in a technique called radioimmunoassay.

To this:

Prior to the development of the EIA/ELISA, the only option for conducting an immunoassay was radioimmunoassay, a technique using radioactively-labeled antigens or antibodies.

The original wording implied that RIA is no longer used, which is inaccurate. 152.133.6.130 21:10, 26 October 2007 (UTC)

WikiProject class rating

This article was automatically assessed because at least one WikiProject had rated the article as start, and the rating on other projects was brought up to start class. BetacommandBot 16:25, 10 November 2007 (UTC)

Applications

I made a couple of changes to the "Applications" section for your perusal. I'm not sure I did the best job of wording the changes, so any comments and/or edits would be appreciated.

1) I added one application of ELISA -- specifically, its use in toxicology.

2) I added a paragraph detailing one method of establishing a cutoff.

--206.194.127.112 (talk) 22:48, 14 May 2008 (UTC)

Hi 206.194.127.112, and thanks for the edit. I'm no expert on ELISA but it looked like a good contribution. I made a small copyedit to make the cutoff interpretation a little easier to follow. Keep up the good work! Adrian J. Hunter^{(talk•contribs)} 05:37, 24 May 2008 (UTC)heerramani

Indirect ELISA

Hi, I'm not expert in ELISA techniques, but I'm affraid the explanation of "indirect ELISA" is a bit confusing. Some times it seams to talk about detection of antibody in serum, but other times is really talking about detection of an antigen in serum:

The steps of the general, "indirect," ELISA for determining serum antibody concentrations are:

1. Apply a sample of known antigen of known concentration to a surface, often the well of a microtiter plate. The antigen is fixed to the surface to render it immobile. Simple adsorption of the protein to the plastic surface is usually sufficient. These samples of known antigen concentrations will constitute a standard curve used to calculate antigen concentrations of unknown samples. Note that the antigen itself may be an antibody.

If I'm not wrong, in Indirect ELISA you don't trate the antibody as an antigen. On the other hand, you adsorve a known amount of antigen in the well, then you coat with serum smaple (containing the unknown amount of antibody to be determined), and finally (after washing) you hybridize with the seconadry antibody for further detection.

I cannot use this as an image because of the copy right, but I think it is very clarifying (belongs to the on-line version of Stryer): http://www.ncbi.nlm.nih.gov/books/bv.fcgi?highlight=ELISA&rid=stryer.figgrp.515

Well, as I said before, I'm not expert in ELISA, so I prefere to expose the problem here rather than modify directly the article. I hope it can help. —Preceding unsigned comment added by Gcarotsans (talk • contribs) 15:30, 2 March 2009 (UTC)

Thanks for pointing this out, Gcarotsans. I'm no expert either, so for now I've tagged the article and brought this to the attention of Wikiproject Molecular and Cellular Biology. Adrian J. Hunter^{(talk•contribs)} 12:15, 3 March 2009 (UTC)

Dear all, please take into account that driven by the willing of an exhaustive description you mixed up all the things...
I have a simple and easier approach:

First group of so called "sandwich" ELISA types (common for all subtypes is the fact that the signal is directly proportional with the concentration of the analyte that you search for):
1. Sandwich ELISA (the antigen that you want to find in a sample is made sandwich between two different antibodies, one immobilized on the well surface and the other enzyme-linked).
2. Bridging ELISA (the antibody that you want to find in a sample is used as a bridge between two antigens, one immobilized on the well surface and the other enzyme-linked).
3. Indirect ELISA (the antibody that you want to find in a sample is used as a bridge between an antigen immobilized on the well surface and an enzyme-linked antibody).

Second group of competitive ELISA (the antigen that you want to find in a sample and the same enzyme-linked-antigen from reagents are in competition for occupying a given slots of antibody immobilized on the well). In this case the signal is inverse proportional with the concentration. ----SimplyJohn---- —Preceding unsigned comment added by 86.121.163.182 (talk) 21:40, 3 March 2009 (UTC)

Please consider adding as the second sentence in Step 4 something like, "The detection antibody might be a natural, unmodified antibody that itself will have to be detected in Step 5, or it might be attached by a covalent bond to an enzyme able to provide a signal later so Step 5 will not be needed." Then, please give a brief explanation or a reference to how this covalent attachment can be made. —Preceding unsigned comment added by Nhy67ygv (talk • contribs) 00:31, 25 March 2009 (UTC)

The section on the sandwich ELISA really states that the capture antibody should be coated on the ELISA plate and does not really discuss the concept of 'sandwich-type' ELISAs as suggested above. I think indirect ELISA should really have it's own section, as most experts would consider these two techniques to be different and to have different applications (one is used to detect antibodies in a sample, the other used to detect antigens). DocLovely (talk) 10:47, 6 October 2020 (UTC)

The "Indirect" section states: "1. Unlabeled antibody is incubated in the presence of its antigen (sample)" ... "5. The sample, that now contains the tagged and bound antibodies,..." To me this does not make sense and is confusing. Also the cited patent states "introducing tagged antibodies into the well". The wiki article does not mention the addition of tagged antibodies. Also why use the words "(un)labeled" and "(un)tagged", which are (often) interchangeable? Do they really mean different things here? If not we should use one term consistently. Since I'm not an expert I don't want to change "1. Unlabeled" to "1. Tagged" right away, but I'd appreciate if an expert could clarify this. --Penschy (talk) 17:52, 11 June 2024 (UTC)

See Also to Lyme disease

ELISA is a critical test for Lyme disease. See Guessing Game which says "The most common Lyme test is called ELISA, which is mostly used among general practitioners not specializing in Lyme disease diagnosis and treatment. However, the ELISA test is considered to be the least sensitive, only detecting the patient’s IgM and IgG levels, which are the body’s reaction to the Lyme bacteria." Simesa (talk) 12:01, 27 June 2009 (UTC)

I'm removing this link because ELISA is a critical test for a wide array of diseases. This is not the place to list all of them. NickCT (talk) 19:12, 15 December 2009 (UTC)

Controversy

I notice that there is no controversy section. I came to Wikipedia to get an objective opinion on this controversial procedure, but there's not even any mention that controversy exists!! Does this mean that no credible criticisms exist? The Quackwatch page on ELISA/ACT was the first result on Google: http://www.quackwatch.com/01QuackeryRelatedTopics/Tests/allergytests.html -- and similar warnings can be found all other the web. I have no opinion one way or the other, but I at least expected to see the controversy mentioned (and hopefully summarized in an unbiased, understandable way). I am definitely a layman in all this stuff, though, so I can't do it myself. --damonhastings

The ELISA is an accurate, accepted method for analysis that has many well-established applications. There is no such controversy about ELISA itself. The link you provide reviews its use to make a controversial claim, such as using an x-ray machine to predict the weather. The x-ray machine is not the problem in that example, but it's application would be. --MartinezMD (talk) 17:21, 6 May 2010 (UTC)

Martinez is right, and the analogy to using an x-ray machine to predict the weather is perfect. Or say the use of gold flakes in a cars gas tank. Neither gold nor gas tanks are controversial, just the quack promoting a fringe connection between the two. ELISA is used to do MYRIAD tasks in biotech, it's no more controversial than western blot or rt-PCR.-Paul 96.41.74.95 (talk) 03:34, 20 December 2013 (UTC)

ELISA vs EIA

What is the really the difference between EIA and ELISA? I think in most ELISA assays the primary antibody is attached to the walls of the microtiter well. Could it be that in EIA the primary antibody is in solution at the beginning of the assay? Benkeboy (talk) 15:13, 26 August 2009 (UTC)

I found an explanation online but I do not quite understand what it means :

"EIA is the more traditional enzyme immunoassay. The technology has been widely used for the analysis of drugs of abuse. It is homogenous in nature meaning that the analysis is performed without any physical separation during the analysis. ELISA is heterogeneous — the microtiter plate is washed before the reaction is allowed to go to completion. In general, ELISA assays may offer greater sensitivity than most EIA procedures."

Benkeboy (talk) 15:19, 26 August 2009 (UTC)

Competitive ELISA

I was given to believe that the competition in this procedure was between the primary antibody, and the secondary antibody, and that it was the concentration of the primary antibody that was under investigation. It is not clear whether the ntibody concntration or the antigen concentration is being investigated in this passage. (Jbonner01 (talk) 13:13, 1 January 2010 (UTC))

Principle

The section is confusing and not at all concise. — Preceding unsigned comment added by 137.43.188.170 (talk) 15:28, 26 April 2012 (UTC)

Indirect ELISA

The washing step is omitted from the description given. It doesn't make sense as it is. — Preceding unsigned comment added by 131.111.129.136 (talk) 15:53, 15 July 2012 (UTC)

Pronunciation

The pronunciations shown (/ɨˈlaɪzə/, /ˌiːˈlaɪzə/) appear to suggest that is it pronounced as a single syllable. Am I misreading it, misinformed, or is the article wrong? Techguy95 (talk) 23:51, 1 May 2014 (UTC)

The article is correct. There are 3 syllables. For help with understanding the transcriptions, you can hover over each symbol for a tooltip (works on desktop, may not work on mobile). You can also click through to the linked Help:IPA for English, which provides more help. Quercus solaris (talk) 21:54, 2 May 2014 (UTC)

ELISA and IGRA

Dear specialists! Please describe differences between ELISA and IGRA. As I understanding ELISA are one of IGRAs (IFN-γ Release Assays)? See PMC2216027 for example. Grumbler eburg (talk) 12:22, 17 January 2019 (UTC)

ELISA and COVID-19

ELISA was identified, in a recent government briefing, as the technique used by Porton Down in its high accuracy COVID-19 antibody tests for population sampling. Although I can't find a transcript of the that part of the briefing, it was also identified as being used by several laboratories in select committee evidence.

David Woolley (talk) 15:37, 8 April 2020 (UTC)

References