Tuesday, July 15, 2008

Lyme antibodies revisited

The ELISA test for Lyme is a good test. The problem is that the cutoff for a positive reaction is set too high. This technology is very good. In this test an enzymatic reaction causes a color change when antibodies react with antigens(Lyme proteins). The amount of color change is transposed into an index. The cutoff point for a positive ELISA was set high when it was discovered that nearly everyone reacted to Lyme on the assay. It was assumed that false positives were occurring. The set point for a positive reaction was decided by a committee in a somewhat arbitrary fashion, who at the time assumed that Lyme disease was rare. Some Lyme antibodies are non-specific and cross reactive. Still the ELISA was supposed to allow for false positives which could be sorted out with the Western Blot test. Positivity of the Western Blot was also arbitrarily set based on X number of bands being positive. This makes no sense. Non-specific bands should be excluded. Bands which specifically react to outer surface proteins of Borrelia burdorferi should be sufficient to make the diagnosis even if only one band is present. Bands 23, 31, 34, 39 and 93 are so specific that a reaction to any of these bands should be considered a positive Lyme test. Indeterminate bands, especially when they occur on several of these specific locations should also suggest a positive test for Lyme. This brings us to the C6 peptide test. This is an ELISA test which correlates with specific reactivity of a very, very specific outer surface protein on the Lyme bacteria. There are virtually no false positives here. The cutoff has been set so high that the test is worthless based on the standards set by the manufacturer. It is interesting that Gary Wormser, the chief critic of chronic Lyme is financially connected to Immunetics, the company which developed this test. There is no reason for any non zero value other than exposure of the immune system to the spirochetes which cause Lyme disease. In fact C6 peptide indices of 0.2 to 0.3 are typically seen in patients who have positive Western Blots even by CDC surveillance criteria. Patients who are treated for Lyme frequently sero-convert. They develop a greater antibody response after antibiotic therapy. By the same token, the C6 peptide index frequently doubles after the patient receives therapy with antibiotics. The standard of a C6 index of greater than 0.9 is ridiculous in my experience. Any level greater that zero is suggestive. Values greater that 0.3 are reliable in confirming the diagnosis. To date there are no published studies comparing C6 index levels to Western Blot positivity. When this data is published it will turn out that C6 peptide index for Lyme disease will be very useful. Is often said we need a better Lyme test. We actually have a better test, we just need to learn how to interpret the results in a meaningful way.


Avalon said...

I was just diagnosed with Lyme 3 weeks ago. I was referred to a Rheumatologist because of ongoing body/muscle aches. he stated he is not certain that I am positive because I only show 3 bands out of 5. My primary said that I am positive by Western Blot, IgG and IgM. The test was repeated twice.

Can you help me understand that?

Lyme report: Montgomery County, MD said...

Look at my blog about Lyme testing written in May or June. It is very complicated. Very few rheumatologist are Lyme literate.

Frosch said...

I agree the C6 is good. I was not WB positive but had a 0.96 C6 titer. I read up about the C6 and all the studies seemed promising, especially that it had few false positives, detects all borrelia, and titers seem to decline with successful treatment. So I got 6 weeks IV followed by 2 months oral abx, and two weeks ago my titer was 0.42. While on IV it went up to 1.2, and I became IgM positive on all three bands through Labcorp. I have only had a few IgG bands, but I have had 2 positive IgM WBs 9 months apart and two positive C6 tests. I also did Igenex once and was IgM positive by their criteria, and also their IFA screening test.

Anyhow, I think the importance of the IgM antibodies are not understood. Many lyme patients seem to produce significant or predominantly IgM antibodies against Lyme-- it is well documented in the literature.

Lyme report: Montgomery County, MD said...

I am sorry I didn't respond to this earlier. It got by me. Great question. Most Lyme patients produce predominantly IgM antibodies. Infectious disease types use this as an argument against Lyme, claiming they are false positives. After all, the standard paradigm suggest that an established infection is associated with IgG antibodies. This easy to explain. Chronic Lyme hides and suppresses the immune system.Lyme spirochetes live in tissues. Antibodies are only produced when the immune cells, B lymphocytes and macrophages encounter germs in body fluid compartments. When antibiotics are given the hidden germs are killed. Pieces of dead Lyme, with their surface protein antigen now exposed to these circulating immune cells. To the immune system it always looks like a new infection. Antibody levels fall quickly with Lyme. IgG antibodies if they are formed quickly disapear to a low or undetectable state. You will notice many "indeterminate" IgG bands on an IgeneX WB test. These are low level reactions which may disappear. The longer someone has chronic Lyme, the less antibodies are found. Ironically, the sickest patients frequently test negative, but may "seroconvert" after antibiotics are given. Clinicians see this response every day. I do not know why the literature does not discus this much.

Frosch said...

It is also clear in the literature that IgM response remains prevalent in those infected with Lyme. And I mean even studies that IDSA folks would consider uncontroversial.

In any case, I'll be relying most on the C6 in my case. I agree with your contention that we already have a better Lyme test in the C6, and like the nuance that anything above 0.1 ought not to be dismissed. The C6 antibodies do not seem to 'hang around' much after they are no longer used, so there is no reason to have a 0.9 threshold.

Lyme report: Montgomery County, MD said...

Most LLMDs don't use the C6 test because the average level in WB positive cases is 0.2 to 0.3. Still, some folks have C6 levels of 2.0 with no WB bands. The C6 test uses the same ELISA technology that most LLMDs have discarded. The original ELISA would be good if labs reported the numeric values like they used to do, rather than an arbitrary cut off which is negative or positive. Some labs are already doing the same for C6 reporting.

Frosch said...

I am not sure most LLMDs have given the C6 a serious look. ELISA is good technology if it is used well. The C6 is a specific recombinant antigen-- you know what you have and that it is relatively pure. The old ELISA is just essentially pulverized Lyme bacteria, so you could get some reactivity only from the flagellin antibodies you might have from something else.

I think it's unacceptable that labs do not report the actual result if they have it. If someone is 0.85, they have the right to know. I have done C6 with Quest, Labcorp and Igenex and they all reported the actual #, even when <0.9.

Samantha said...

My adult daughter appears to have chronic lyme and has been treated for years. The last 6 months she has declined and so her LLMD retested her and she is CDC positive. (IgM 23,41 and IgG 18,23,28,39,41,66,93) Question her C6 came back high 3.26. Does this indicate the level of infection? She is startin on bigger gun orals and if not better back to IV's.



Lyme report: Montgomery County, MD said...

The level of C6 may indicate disease activity. It does frequently improve with treatment.

The only reliable test for disease activity is an assessment of symptoms and perhaps an appropriate physical exam.

Lab results in general should not be used to guide therapy.

Powers said...

I have a question for anyone who is educated on the C6 Peptide test and can speak to its specificity, please. I tested positive on the C6peptide, with a very high titer of 4.1. However, I did not test positive on the Western Blot. My physician felt strongly that it was Lyme based on my symptoms and the high titer, so she treated me with a month of oral doxy. Within a week of treatment, my gastrointestinal issues that have been plaguing me for years discipated. I feel much better, however am haunted by the possibility that something else is going on in my body and being left untreated, and that the positive C6 may have been a false one. Any information you can provide to help me better understand these results would be very much appreciated.

Unknown said...
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Unknown said...

So, according to what you said about chronic lyme wouldn't the antigens be circulating all the time so it would show up on the nano-trap antigen test? This would be true even if the body isn't making IgG antibodies because the bacteria are not circulating? The ospA would still be circulating to be detected on blood?

You said:
When antibiotics are given the hidden germs are killed. Pieces of dead Lyme, with their surface protein antigen now exposed to these circulating immune cells.

So, if somebody tests positive on IgM band 31 right now and this is supposed to be specific and indicative for chronic lyme then one would expect for that person to test positive for the antigen ospA on the nano-trap antigen test if that is the true cause of band 31 IgM, right?

If the ospA super sensitive nano-trap antigen test is negative, this would seem to contradict the theory that band 31 IgM is a true positive.

It will be interesting to see what happens to confirm this theory.

The antibiotic provocation should not be needed to detect ospA antigens with the antigen test. Perhaps there are enough ospA antigens to show up in blood, but not enough to convert to IgG?

What do you think?

Unknown said...

I don't know what the units are for the c6 in humans, but the c6 test for dogs and horses says anything above 30U/ml is positive. I think that is .3 on the human test. But, why would somebody be producing these at all if they didn't have it? Are tests just not able to filter out the noise below a certain point?

The nano trap antigen test in 2015 will include VlsE outer surface protein (among others) which the C6 segment is part of, so we should be able to test for C6 antigen directly in humans in 2015.

The C6 is the sixth invariable region of the VlsE protein, a 25-amino-acid peptide termed "C6". The C6 ELISA is an antibody test that tests for antibodies to the C6 antigen. However nano-trap antigen tests test for the C6, ospA etc antigens directly. We should be able to test for ospA in 2014 and other antigens in 2015.

There was not a penny of federal funding to develop these human tests (not to mention a cure). Patients had to lobby the state of Virginia to fund a small amount of money for a small company.

Meanwhile trillions are being spent to find a cure for "Global Warmings".

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Unknown said...

My Elisa shows a result of .34 which technically isn't high enough to be considered "positive." My Western Blot shows only Band 39 IgM. I have had many symptoms for many years. Would you consider an Elisa result of .34 to be confirmatory of Lyme?

K10 said...

I am trying to understand my recent lab results. I have been reading that they are considering replacing the the two tier system with the C6. My results on the C6 test was positive with a high range score of 4.51. My blot test was negative. How can these be so different? Which one is the false negative? I check all the boxes concern Lyme including having had a circular ring rash. The only issue is the negative blot so my PCP says I am negative then. I am searching for a new doctor but at this point I want to be well inform and know how to advocate for myself.