Explaining Borelliosis (Lyme) Western Blot Tests

The Western blot is a type of test that is conducted for detection of
Borreliosis (Lyme), but is also used to test for infections other than
Borreliosis.

Borreliosis is a more accurate name than Lyme disease for this infection.
Several different Borrelia may cause a similar clinical pattern in this disease.

There is no universal agreement on what defines a positive Western blot.

Good laboratories use different criteria to interpret borreliosis blots. At the
1999 international borreliosis and tick-borne infection conference, Sam D****,
M.D. Lectured.

Dr. D**** is a full professor of Infectious Disease at Boston University School
Of Medicine. He said that if a patient has just one borreliosis-associated
Antibody on their Western blot, you may assume they have borreliosis. Richard H*****, M.D. Said the same thing in his lecture, at that same conference.

Research I presented in 1998 involving over 400 borreliosis patients, showed an 87% response rate to antibiotics. This was if they had one Borreliosis-associated antibody on their blot.

So if there is enough suspicion that Lyme borreliosis is the cause of a
Patient's' symptoms, so much so that a Western blot is ordered, then if only one Borreliosis-associated antibody is found, it is significant!

Medical literature is replete with statements about false positive test results
For Lyme borreliosis. Since 1988, I have diagnosed and treated well over 600
Borreliosis patients. Only 2 of those patients with a positive borreliosis test
Did not respond to antibiotics. This is a 99% success rate!

So in the trenches of day-to-day medical practice, false positive borreliosis
Tests are not an issue. In retrospect, those 2 patients that did not respond to
Antibiotics may have also had babesiosis.

In my practice, many borreliosis patients also have babesiosis, another
Tick-borne infection that causes the same symptoms as Lyme borreliosis.

Babesiosis is caused by a protozoa, which is a different germ type than a
Bacteria, virus, fungus or yeast.

The placebo effect would not explain a 99% response rate. Those borreliosis
Associated antibodies should not be there, in patients with symptoms.

A placebo is like a sugar pill, that has no effect. A placebo effect occurs
Because patients believe in the pill they are taking, even though it is a sugar
Pill. The human mind causes the response. Placebo effects should more likely be About 20-30%, not a 99% response rate.

False negative test results are the real problem in diagnosing borreliosis.
Research has shown that you have to do the right test (the Western blot), done. At the right laboratory (one that specializes in testing borreliosis), and done The correct way (shipped express delivery early in the week).

The right test to screen for borreliosis is the Western blot. Research I
Presented in Bologna , Italy in 1994 at the international borreliosis conference
Showed this.

Other screening tests, such as the IFA, EIA, ELISA, and PCR DNA probe were often Negative when the Western Blot was positive!

Other doctors like myself who diagnose and treat a lot of borreliosis patients,
Go straight to the Western blot as their screening test.

Medical articles abound stating that it is best to do a screening test, such as
An ELISA, and if it is positive, then confirm it with a Western blot.

But the ELISA is often negative when the Western blot is positive so, the right
Test is the Western blot.

It lets you see exactly which antibodies are present. The "right laboratory"
Means one that specializes in borreliosis testing.

In the past, I have done head to head comparisons with 3 different regular labs.Western blots were drawn and sent on the same day to 2 different labs.

The labs that specialize in borreliosis testing typically found
Borrelia-associated antibodies, that the regular laboratories missed.

If these specialty labs find a borrelia antibody, I trust it to be significant,
Because patients respond to antibiotics.

You get what you pay for, so use a lab that specializes in borreliosis. The
Right way to process the Western blot specimen means for the blood to be drawn
and express mailed early in the week.

Research shows the borrelia antibodies have the potential to clump together,
resulting in false negative test results. So far, unclumping has not been
practical for laboratories to do.

The fresher the specimen, the more accurate the test results. Patients at our
office are scheduled Monday, Tuesday, or Wednesday if testing is to be done.

This way, express shipping will assure that the specimen does not spend the
weekend sitting at the post office. This is the right way to test and ship
borreliosis specimens.

Western blots look for antibodies. These antibodies are made by your immune
system. In this case, the antibodies are made to fight against different parts
of the Lyme bacteria, which is called Borrelia burgdorferi, and other Borrelia
species.

In other words, your immune system does not make one big antibody against the whole bacteria. So, when you see a number on a borreliosis Western blot, it corresponds to a specific part of the bacteria.

Compare it to the old story of different blind people touching an elephant.
Based on the part of the elephant each one touched, each person had their own perception. Likewise, the antibodies attach to different and specific parts of Borrelia burgdorferi.

Numbers on Western blots correspond to weights. Kilodaltons (kDa) are the units used for these microscopic weights. Think of it like pounds or ounces. An 18 kDa antibody weighs 18 kilodaltons.

To do a Western blot, thin gel strips are impregnated with the various parts of
Borrelia burgdorferi. Each of the numbers, 18 through 93, on the test result
form, is a part of the bacteria.

Blood is made up of red blood cells and serum; Spinning blood in a centrifuge
separates serum from red blood cells and other things, like white blood cells
and platelets.

Serum contains antibodies made by the immune system. Electricity is used to push the serum through the thin gel strips for the Western blot.

If there are any antibodies against parts of Borrelia burgdorferi present in
your serum, and these parts are impregnated on the strip, the antibody will
complex (bind) to that part.

When antibodies form a complex, it is called an antigen-antibody complex.
Anything foreign in the body is an antigen, such as a ragweed pollen particle,
germ, cancer, and even a splinter.

In the case of borreliosis, the various parts of Borrelia burgdorferi are all
antigens. Though each antigen is different, they all come from the same
bacteria. So all the numbers that are positive on the test report are due to
antigen-antibody complexes.

If enough of the complexes are formed, eventually it may be seen with the naked eye as a dark band. - Band intensity reflects how dark or wide it is.
Controversy exists about band intensity.

Many would say the " +/-" equivocal [" IND "] bands are not significant. The
problem I have with that, is that there are "-" negative bands. The lab has no
trouble calling some bands negative. So they must be seeing something when they
put "+/-" at some bands.

The only thing that makes sense, is that there is a little bit of that antibody
present in your serum. If the "+/-" equivocal is reported on the borrelia
associated bands, it is usually significant, in my clinical experience. This is
a strong clue that I am on the right track.

Instead of ignoring these, they should be a red flag to keep pursuing a
laboratory diagnosis. Giving patients 4 weeks of antibiotics (usually
tetracycline, 500 mg, 3 times a day), will convert a negative or equivocal
Western blot to positive in about 36% of cases.

As mentioned, if these positive blots are found by specialty labs, over 99% of
those patients will respond to antibiotics.

Sometimes multiple antibiotics have to be tried before the patient feels better.
Antibiotics may actually help with the laboratory diagnosis. But patients need
to be off antibiotics about 10 to 14 days before the Western blot is repeated.
This sounds like a contradiction.

Antibiotics may help convert the test to positive, but patients need to be off
antibiotics when the specimen is drawn.

It is well documented in medical literature that the presence of antibiotics may
cause false negative borreliosis testing. Therefore, your system should be free of all antibiotics for an accurate blot result.

When the Lyme borrelia are alive, they are geniuses at avoiding the immune
system. They may do things like go inside your white blood cells, and come out enclosed by the cell membrane of your own white blood cells! This may partly explain why antibodies against Borrelia burgdorferi are often not found when patients are tested.

What may happen when patients are given 4 weeks of tetracycline (or other
antibiotics) is that some of the bacteria die. When Borrelia burgdorferi dies,
it is less efficient at avoiding the immune system.

That's when antibodies may be formed against Borrelia burgdorferi, converting the negative or equivocal Western blot to positive, in about 36% of cases.

If a borreliosis Western blot is going to be positive, it is usually the first
one that is positive. The second blot is the next most likely to be positive,
and so on, until the fifth blot.

After that, the curve levels off for conversion to positive. This is based on
research I presented in Bologna , Italy in 1994. Some patients had
borrelia-associated antibodies finally show on their tenth Western blot! Two
Western blots from a reliable lab usually gives the answer.

If a third test is needed, a Lyme Urine Antigen Test (LUAT) is done instead of a third Western blot. Positive LUATs correspond very highly to patients getting
better with antibiotics.

False positive LUATs have not been a problem in my practice. The LUAT finds the actual antigen (Borrelia burgdorferi itself), so arguably it should be the test of choice, but the Western blot is rn6re widely accepted, even though it looks for the antibodies against Borrelia burgdorferi.

The presence of antibodies are indirect evidence of an infection, not direct
evidence like shown in the LUAT. On the Western blot test result form, please
note what is "considered positive" and "considered equivocal." Equivocal is
another way of saying suspicious or almost positive.

Below this are the ASTPHLD/CDC recommendations. The CDC stands for the Center for Disease Control. I have been in attendance at the international borreliosis conferences when the CDC said their recommendations are for disease surveillance, not day-to-day clinical medical practice. I am not in the business of disease surveillance. My job is to try to help sick people.

The CDC recommendations do not include the 31 and 34 Kda bands of the blot test. These two bands correspond to outer surface proteins A and B respectively (ospA and ospB).

In the world of borreliosis, these are two of the classic hallmark Lyme
antibodies. But the CDC does not even have them in their recommendations.

You may see why I and other borreliosis clinicians do not agree with using the
CDC criteria in everyday medical practice. Other bacteria besides Borrelia
burgdorferi may produce the 45, 58, 66, and 73 kDa bands.

These bands may be produced by Borrelia burgdorferi, but are not nearly as
specifically associated with Lyme borreliosis as the starred bands. These
starred bands are classic hallmark borrelia-associated antigen-antibody
complexes.

An example of the CDC's criteria of a blot test, is if a patient has the band
pattern of 41, 45, 58, 66, and 93, the CDC would call it positive. But if a
patient has a 23-25, 31, 34, and 39 band pattern, they would call it negative.

This is despite the fact that this second pattern of antigen-antibody complex
bands is much more specifically associated with Borrelia burgdorferi than the
first pattern.

As you can see, borreliosis is very controversial. It would be alarming if I was
the only clinician who thought that the CDC recommendations should not be used for day-to day medical practice.

Many borrelia clinicians do not use the CDC criteria. This is obvious by the
fact that the IgX laboratory uses different criteria for positive. Again, in my
opinion and others', even one borrelia-associated antibody is significant, if
symptoms exist.

The classic triad of symptoms for borreliosis is fatigue (tiredness,
exhaustion), musculoskeletal pain (joints, muscles, back, neck, headache), and cognitive problems (memory loss, trouble concentrating, difficulty remembering what you read, depression, disorientation, getting lost).

But there are about 100 symptoms on the borreliosis questionnaire I use.
Borreliosis may mimic or imitate virtually any disease.

Patients often tell me that other physicians they have seen use the CDC
recommendations. This is unfortunate, in my opinion, since these physicians are not in the business of disease surveillance, like the CDC is.

But I am biased. After seeing patients with borreliosis since 1988, attending
many conferences, talking with experts, and doing research on borreliosis
testing, there is absolutely no question in my mind that physicians need to not
blindly accept any recommendations.

One of my hopes is that doctors will someday realize that this controversy is a
signal for them to search for the truth. Why is there such conflict in this very
"political" disease if there is not substance for disagreement? Both IgG and IgM Western blots should be done for borreliosis.

With most infections, your immune system first forms IgM antibodies, then in
about 2 to 4 weeks, you see IgG antibodies. In some infections, IgG antibodies may be detectable for years.

Because Borrelia burgdorferi is a chronic persistent infection that may last for
decades, you would think patients with chronic symptoms would have positive IgG Western blots.

But actually, more IgM blots are positive in chronic borreliosis than IgG. Every
time Borrelia burgdorferi reproduces itself, it may stimulate the immune system to form new IgM antibodies.

Some patients have both IgG and IgM blots positive. But if either the IgG or IgM blot is positive, overall it is a positive result.

Response to antibiotics is the same if either is positive, or both. Some
antibodies against the borrelia are given more significance if they are IgG
versus IgM, or vice versa.

Since this is a chronic persistent infection, this does not make a lot of sense
to me. A newly formed Borrelia burgdorferi should have the same antigen parts as the previous bacteria that produced it.

But anyway, from my clinical experience, these borrelia associated bands usually predict a clinical change in symptoms with antibiotics, regardless of whether they are IgG or IgM. In regard to the outer surface proteins, think of it like the skin of a human.

On the outer surface of the Lyme bacteria are various proteins. As they have
been discovered, they have been assigned letters, such as outer surface proteins A, B, and C.

The following is a brief explanation of the test results. Again, each band is an
antigen complexed (bound together) with an antibody made by the immune system, specifically for that antigen (part) of Borrelia burgdorferi.

18: An outer surface protein.

22: Possibly a variant of outer surface protein C.

23-25: Outer surface protein C (osp C).

28: An outer surface protein.

30: Possibly a variant of outer surface protein A.

31: Outer surface protein A (osp A). 34: Outer surface protein B (osp B).

37: Unknown, but it is in the medical literature that it is a
borrelia-associated antibody. Other labs consider it significant.

39: Unknown what this antigen is, but based on research at the National
Institute of Health (NIH), other Borrelia (such as Borrelia recurrentis that
causes relapsing fever), do not even have the genetics to code for the 39 kDa
antigen, much less produce it. It is the most specific antibody for borreliosis
of all.

41: Flagella or tail. This is how Borrelia burgdorferi moves around, by moving
the flagella. Many bacteria have flagella. This is the most common borreliosis
antibody.

45: Heat shock protein. This helps the bacteria survive fever. The only bacteria in the world that does not have heat shock proteins is Treponema pallidum, the cause of syphilis.

58: Heat shock protein.

66: Heat shock protein. This is the second most common borrelia antibody.

73: Heat shock protein.

83: This is the DNA or genetic material of Borrelia burgdorferi. It is the same
thing as the 93, based upon the medical literature. But laboratories vary in
assigning significance to the 83 versus the 93.

93: The DNA or genetic material of Borrelia burgdorferi.

In my clinical experience, if a patient has symptoms suspicious for borreliosis,
and has one or more of the following bands, there is a very high probability the patient has borreliosis.

These bands are 18, 22, 23-25, 28, 30, 31, 34, 37, 39, 41, 83, and 93.

This is true regardless of whether it is IgG or IgM.. But again, there is no
universal agreement on the significance of these bands. Betina Wilska, M.D. from Germany is one of the world's experts on outer surface protein A (31 kDa).

At the international borreliosis conference in Vancouver , British Columbia , I
asked her personally about the 30 kDa band. She told me it was the same as the 31 kDa band (osp A).

When you have the opportunity to talk to borreliosis experts, this helps in
assigning significance to findings, on an imperfect test. As a medical doctor, I
am stating all of this with no axe to grind, no professorship to protect, and no
preset opinions. Patients, personal research, and conferences have helped me interpret the borreliosis medical literature in regard to testing.

Nobody would like to have available a bullet-proof, 100% reliable Lyme
borreliosis test more than I would. But we must use what is currently available. I always welcome second opinions.

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Here is his update written sometime around 2005.

When physicians do consider borreliosis, they often start with a screening test
such as an EIA, ELISA, IFA or PCR-DNA probe. If the initial screening test is
negative, many physicians tell patients they do not have Lyme borreliosis and
the testing is stopped right there.

Screening tests that are positive are often followed by a test called the
Western blot. The blot is a "confirmatory" test, as opposed to a screening test.

(Blots are performed for other infection -- it is a type of test, not a test
uniquely for the Lyme bacteria.)

Western blots are accomplished by breaking the Borrelia burgdorferi into pieces,
and those parts of the Lyme bacteria are then embedded in a gel.

Electricity is used to push antibodies made by the immune system through the
gel. Antibodies that are made to attach to certain parts of the Lyme bacteria
will bind to those exact parts that are embedded in the gel.

When the antibodies bind to the parts of the bacteria, a black band is formed,
which is then interpreted as +/-, +, ++ or +++ depending upon the intensity or
darkness of the band.

Each part of the Lyme bacteria weighs a certain amount