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The MastAttack 107: The Layperson’s Guide to Understanding Mast Cell Diseases, Part 13

I have answered the 107 questions I have been asked most in the last four years. No jargon. No terminology. Just answers.

21. Why do people care so much about diagnostic criteria?
• Historically speaking, the medical establishment tends to draw very explicit borders around diagnoses. There are several reasons for this.
• It is partly to help diagnose things correctly. There are thousands and thousands of diseases and disease states. The most effective way of getting as many people as possible correctly diagnosed with a disease is to define what that disease is and how you diagnose it. That doesn’t mean that every person who has this disease will always be diagnosed correctly. It also doesn’t mean that every person who doesn’t have this disease will be diagnosed with something else. It just means that this is the best way to diagnose the largest point of people all over the place.
• It is also to strength any research done around these diagnoses. As a scientist, who has to operate within the trappings of specific diagnoses with specific criteria, it is 100% necessary for me to do my job well.
• We have to know that all the patients in a study meet the same criteria. It’s not enough for their doctor to give them a diagnosis because they think that’s what they have even if they don’t meet the criteria. Let’s look at this a little more closely below, under the heading “Blue Disease.”
• The bottom line is that diagnostic criteria is the foundational bedrock of the Western medicine establishment (and some Eastern traditions as well).
Diagnostic criteria also help determine what insurance companies will pay for. If you are a provider caught saying a bunch of patients have a diagnosis that they don’t have, you can be charged with insurance fraud. That can carry significant penalties including fines, loss of license and even prison time.
• Furthermore, if a doctor is caught misdocumenting diagnosis, insurance companies will crack down on patients with the same diagnosis in other places, making it harder for everyone to get treatment. There have been situations in recent history where patients getting a very expensive treatment were required to stop treatment to prove that they needed it since doctors were prescribing it for many other conditions without documenting it correctly.
• The last reasons why everyone cares about diagnostic criteria are related more to the experiences of patients within this community. Most of us have been misdiagnosed more than once. It can really complicate things and it can endanger people. It can also really scare people, too.
• Finally, most of us in this community have been lied to someone impersonating a rare patient at least once and usually more. It is exhausting and insulting.
• I want to be very clear that the reason a lot of people get stuck on diagnostic criteria is NOT because people who don’t meet one or the other set are not deserving of treatment or are not as sick. That is not the case at all.

Blue Disease:

• Let’s say that I am running a study on a disease called Blue Disease. Blue Disease is a condition that strikes people on their 25th birthday. On this day, people with this disease just wake up completely blue. They are never not blue again. I am interested in Blue Disease and so I design a study for it.
• In order to fund my study, I have to get grant funding. This money may be from a private foundation or a university or the government. I have to convince them to care about Blue Disease. More importantly, I have to convince them that the money they give me will be used intelligently and not wasted.
• Let’s say that I let in 100 people who all tell me they have Blue Disease. They are all blue. They all are older then 25. I let them in to my study to research a medicine to treat this disease.
• At the end of my study, I have found that if I give most of them a medicine called anti-Blue, their blue goes away. There is gladness and rejoicing. I find that 90 out of 100 respond to the medicine. Hooray! That’s a 90% success rate.
• Except then I find out that not all of those people actually had Blue Disease. Some of them turned blue before their 25th birthday. Some of them started purple, then became blue, then green. And so instead of having a 90% success rate for Blue Disease, we find that it’s much less effective than 90% for Blue Disease. We know that it has helped some other people not be blue but we don’t even know what disease they have. And I am in a hell of pickle as a researcher because I don’t know what these data mean.
• Because the medication seems not very effective for Blue Disease, it doesn’t get approved or prescribed to people who have Blue Disease.
• Because my study was not controlled enough, no one wants to give me any more money to research this disease. In certain situations, I could actually have to pay back the money, would almost certainly lose my job, and could be prosecuted because I have an ethical obligation to only research the disease I say I will research in a study.

For more detailed reading, please visit these posts:

The Provider Primer Series: Mast cell activation syndrome (MCAS)

The Provider Primer Series: Cutaneous Mastocytosis/ Mastocytosis in the Skin

The Provider Primer Series: Diagnosis and natural history of systemic mastocytosis (ISM, SSM, ASM)

The Provider Primer Series: Diagnosis and natural history of systemic mastocytosis (SM-AHD, MCL, MCS)

Chromogranin A

Chromogranin A is a protein secreted in several environments. While it is primarily released in the adrenal medulla with catecholamines (norepinephrine, epinephrine, dopamine, and others), CgA is often found stored in the granules of endocrine cells in the GI tract. CgA is the precursor molecule for several active molecules. Vasostatin-1 and -2 are involved in regulation of various effects of the cardiovascular system, including blood pressure and stroke volume, by opposing the action of catecholamines. Catestatin decreases release of catecholamines. Pancreastatin decreases insulin secretion. A number of other molecules are also derived from CgA.

Chromogranin A and its derivatives are biomarkers for several conditions. 60-80% of neuroendocrine tumor patients demonstrated elevated chromogranin A. A connection with Alzheimer’s disease has recently been reported. Rheumatoid arthritis and lupus patients may have elevated CgA as a result of increased tumor necrosis factor. Various forms of cancer, kidney disease, and elevated cortisol can also impact chromogranin A level.

Elevated CgA has also been linked to a number of inflammatory GI conditions. 30-50% of IBD patients with active disease have elevated serum CgA. In ulcerative colitis, fecal chromogranins were elevated but not correlated with disease activity. Conflicting results have been seen in patients with Crohn’s disease. Some studies have reported an increased amount of CgA containing cells in patients with IBS.

There are a number of methods for quantifying chromogranin A. Proton pump inhibitors and H2 antihistamines can yield false positive results. A study compared several commercial kits for measuring chromogranin A and found that the radioimmunoassay (RIA) kit was most likely to be accurate with a sensitivity of 93% and specificity of 85%. This means that 93% of the time, this kit properly identified patients with high CgA as having high CgA, while 85% of the time, it properly identified patients with normal CgA as having normal CgA. Currently, there are multiple test methods for quantifying serum and plasma CgA with no central standardization.

Chromogranin A is a constituent of granules in rat mast cells. Tumor necrosis factor is a mediator released by mast cells and may also influence the levels of chromogranin A in mast cell patients. One study found that 31.5% of patients with mast cell activation disease (in a cohort mostly composed of MCAS patients) demonstrated elevation of serum CgA. This same study concluded that plasma heparin and 24 urine testing for prostaglandin D2 and 9a,11b-prostaglandin F2 were the most sensitive markers for mast cell activation with other mediators being less effective.


Gut P, et al. (2016) Chromogranin A – unspecific neuroendocrine marker. Clinical utility and potential diagnostic pitfalls. Arch Med Sci, 12(1): 1-9.

Wernersson S, Pejler G. (2014). Mast cell secretory granules: armed for battle. Nature Reviews Immunology, 14: 478-494.

D’Amico MA, et al. (2014) Biological function and clinical relevance of chromogranin A and derived peptides. Endocrin Connect, 3(2):R45-54.

Mazzawi T, et al. (2015) Increased chromogranin A cell density in large intestine of patients with irritable bowel syndrome after receiving dietary guidance. Gastroenterology Research and Practice, Article ID 823897.

Zenker N, Afrin LB. (2015) Utilities of various mast cell mediators in diagnosis mast cell activation syndrome. Blood, 126:5174.

Massironi S, et al. (2016). Chromogranin A and other enteroendocrine markers in inflammatory bowel disease. Neuropeptides, xxx, xxx-xxx.

The Provider Primer Series: Natural history of SM-AHD, MCL and MCS

Natural history of systemic mastocytosis with associated hematologic disease (SM-AHD):

  • SM-AHD is defined by systemic mastocytosis in the presence of another clonal hematologic disease. SM-AHD is thought to comprise 30-40% of all mastocytosis cases[i].
  • In about 90% of cases, the associated blood disorder is a myeloid neoplasm such as myelodysplastic syndrome, myeloid leukemias, or myeloproliferative diseases such as polycythemia vera or essential thrombocythemia[i] . Janus kinase 2 (JAK2) V617F mutation, which has a known association with myeloproliferative neoplastic conditions such as essential thrombocythemia and polycythemia vera, is sometimes present in SM-AHD patients[vii].
  • In this condition, SM and the other blood disorder are treated as separate entities as if they did not co-occur[i]. The conditions are synchronous and the associated hematologic disease does not occur secondarily to SM or treatment thereof. Prognosis in SM-AHD depends almost exclusively upon the associated hematologic concern. In multiple studies, fatalities are reported as result of associated malignancies[ii].
  • Myeloid neoplasms are the most common AHD, including chronic myelomonocytic leukemia or other leukemias, myelodysplastic syndrome, or myeloproliferative diseases[i].
  • In a 138 patient cohort: about 1/3 demonstrated Hgb <100 g/L and platelets<100×109/L; 51% had elevated white cell count; 31% demonstrated frank eosinophilia <1.5×109/L[vii].
  • SM-AHD patients are at increased risk of leukemic transformation relative to other forms of systemic mastocytosis (excluding mast cell leukemia) with a frequency of 14% in a 138 patient cohort[vii].
  • The SM aspect of SM-AHD is diagnosed and staged according to the SM diagnostic algorithm. It is therefore possible for a patient with SM-AHD to have mast cell leukemia or any other subtype of SM[vii].

Natural history of mast cell leukemia (MCL):

Table 1: Diagnostic criteria for mast cell leukemia[iii] 
Meets criteria for systemic mastocytosis Mast cells compromise 20% of all nucleated cells in blood smears


Table 2: C findings present in acute MCL[iii] 


C findings One or more cytopenias (absolute neutrophil count <1000/µl; Hemoglobin <10g/dl; platelets <100000/µl) Hepatomegaly with ascites, elevated liver enzymes with or without portal hypertension Splenomegaly with hypersplenism Malabsorption evidenced by low albumin and weight loss Large osteolysis and/or severe osteoporosis and pathologic fractures (2 or more fractures as direct result of mast cell activity)


  • Mast cell leukemia is defined by SM where ≥20% nucleated cells in marrow are mast cells. In leukemic variant, >10% of nucleated cells in blood are mast cells; in aleukemic variant, there are <10% mast cells[iii].
  • MCL can occur de novo or from a previous mast cell neoplastic condition such as aggressive systemic mastocytosis or mast cell sarcoma[iii].
  • CKIT D816V mutation is less common in MCL than in other forms of systemic mastocytosis (50-80%). Some patients have mutations elsewhere in the coding regions of CKIT or a non-D816V mutation at CKIT codon 816. An unusual feature of MCL is that when the disease progresses quickly, the patient may lose positivity for the D816V mutation[iv].
  • MCL patients do not typically demonstrate mastocytosis in the skin[iii].
  • In the absence of C findings, some MCL patients have stable disease without markers of progression. This is referred to as chronic MCL[iii] .
  • >90% mature mast cells is a positive prognostic indicator. Presence of mostly immature mast cells is associated with more aggressive disease[iii] .
  • Acute MCL rapidly causes catastrophic organ damage. Median survival in acute cases is six months, though some MCL patients live for years through the use of newer targeted therapies[iv].
  • Hematopoietic stem cell transplant (HSCT) is an experimental option for MCL patients. In one study, overall survival at the three year mark was 17% (2 of 12 patients)[viii].

Natural history of mast cell sarcoma:

  • Mast cell sarcoma is an exceedingly rare tumor with high grade cytology that can present in a variety of tissues[v].
  • Mast cells that comprise the tumor resemble neither morphologically normal mast cells or spindled cells often seen in SM. In mast cell sarcoma, mast cells are often bilobed and multinucleated tumor cells have been reported. Of note, mast cells compromising the sarcoma often express CD30[iv].
  • Mast cell sarcomas often have neither CKIT D816V mutation nor mutations elsewhere in CKIT coding regions[vi].
  • Mast cell sarcomas often induce only local symptoms at the time of diagnosis but systemic involvement rapidly follows. Mast cell sarcoma may progress to mast cell leukemia. Median survival is 12 months[vi].


[i] Gotlib J. (2013). Approach to the diagnosis and management of mastocytosis. The Hematologist, 10(1). Retrieved from:

[ii] Wang SA. (2013). Systemic mastocytosis with associated clonal hematologic non-mast cell lineage disease (SM-AHNMD): clinical significance and comparison of chromosomal abnormalities in SM and AHNMD components. Am J Hematol, 88(3), 219-224.

[iii] Valent P, et al. (2014). Refined diagnostic criteria and classification of mast cell leukemia (MCL) and myelomastocytic leukemia (MML): a consensus proposal. Ann Oncol, 25(9), 1691-1700.

[iv] Youk J. (2016). A scientific treatment approach for acute mast cell leukemia: using a strategy based on next-generation sequencing data. Blood Res, 51(1), 17-22.

[v] Ryan RJH, et al. (2013). Mast cell sarcoma: a rare and potentially underreecognized diagnostic eneity with specific therapeutic implications. Modern Pathology, 26, 533-543.

[vi] Georgin-Lavialle S, et al. (2013). Mast cell sarcoma: a rare and aggressive entity – report of two cases and review of the literature. JCO, 31(6), e50-e57.

[vii] Lim KH, et al. (2009). Systemic mastocytosis in 342 consecutive adults: survival studies and prognostic factors. Blood, 113(23), 5727-5736.

[viii] Ustun C, et al. (2014). Hematopoietic stem-cell transplantation for advanced systemic mastocytosis. J Clin Oncol, 32(29), 3264-3274.

Lyme Disease: Chronic Lyme (part 5)

The number of patients diagnosed with negative serology Lyme seems to be ever growing. These patients largely have nonspecific symptoms and no obvious explanation for them. They may or may not remember a previous tick bite. Patients in these groups repeatedly test negative for Lyme in various validated tests. This population account for a much higher number of Lyme diagnoses than PTLDS.

Negative serology Lyme patients are diagnosed by medical professionals who believe that they have active, ongoing Borrelia infections despite negative tests. They believe that while taking continuous antibiotics, Borrelia is driven to form hardy structures that can survive adverse conditions (sort of like how molds can form spores that survive for thousands of years.) When the antibiotics are discontinued, patients once again become symptomatic. They believe this is because the Borrelia convert back to infectious forms.

Furthermore, negative tests are ignored for these patients on the basis that Lyme diagnostics are not good. And they’re not.

There is a lot going on here. So let’s look at the data.

I discussed testing at great length in one of the other posts. They function well in a certain time period for a certain group of people. Importantly, those people likely constitute the majority (but not all) of true Borrelia cases in the US. The data on this is currently being generated in a very large, three prong study by the CDC.

When most people get an infection, they generate IgM and IgG antibodies that last for some time. IgM concentrations decrease after about a year (this varies a lot depending on what the infecting organism is, so I’m being general here), but IgG concentrations often persist for years, sometimes even decades. This is why people who got the chicken pox as a kid are unlikely to get it when their kid gets it thirty years later. Your body has really fascinating mechanisms for remembering pathogens for a long time.

But exactly how long do those IgG antibodies last? It’s hard to say. And it’s especially hard to say for this population because they repeatedly test negative, sometimes immediately after onset of symptoms. So I thought the best way to figure this out would be look at how long people with serology positive Lyme disease (positive for IgM or IgG) test positive.

Importantly, the papers that look at this specific issue were mostly written before the CDC recommendation for 2-tier testing (ELISA and blot.) It is really important when you are comparing data sets on a particular topic that they use the same criteria. So while I did read them, I did not feel they were an accurate representation of what I was looking for.

A 2006 paper (Glatz 2006) looked at the IgG and IgM antibodies to Borrelia burgdorferi in 113 patients who had had the EM rash. They analyzed samples taken before treatment began (using standard, short term antibiotic treatment) and samples taken at least one year after treatment concluded. 12% of patients were positive for IgM before and after treatment; 11% were positive for IgG before and after treatment. 56% were negative for IgG before and after treatment; 42% were negative for IgM before and after treatment. 43% were positive for IgM before treatment and later became negative; 30% were positive for IgG before treatment and later became negative.

This study used ELISA testing, which is not likely to miss a positive antibody response. Before therapy, IgG and IgM tests are negative in about half of patients with the EM rash due to the time lapse in the way your body makes antibodies. Also, sometimes all the Borrelia spirochetes stay in the skin at the site of the rash, and your body is less likely to make antibodies to things that don’t actually go inside your body. So the initial negative is not as surprising as the fact that many patients never seroconvert (or that the tests never detect this seroconversion.) Importantly, this study did NOT find that persistent positivity correlated with poor outcome. However, it did find that patients who were persistently positive were more likely to have larger EM rashes or that those rashes lasted longer.

A 2014 paper (Fallon 2014) looked at the accuracy of Lyme tests (the CDC recommended tests and others done by Lyme labs) by using patient samples from two previous studies. All of these patients had tested positive, some by ELISA and some by IgG western blot. The first group of 37 was recruited from 1999-2005; the second of 11 was recruited from 2005-2007. In the Lyme patients, 62.2-67.6% was positive by ELISA; 2.7-43.2% were positive by IgM western blot; 43.2-56.8% were positive by IgG western blot; and 37.8-48.6% were positive by the two tier CDC recommended testing. (More on this tomorrow.)

The samples in these papers were from the time period surrounding treatment, which in some cases was ten years earlier. Antibodies are very sensitive. They are easily influenced and can deteriorate if not stored or handled correctly.

A 2001 paper looked at the current serum antibody response to 79 patients who had previously been serology positive for IgM or IgG 10-20 years earlier. None of these patients had any ongoing symptoms or signs of active Lyme disease. This study used the CDC recommended two-tier process. Among patients who had only had early disease, 10% were still IgM positive and 25% were still IgG positive. Patients who had had Lyme arthritis, 15% were still IgM positive and 62% were still IgG positive.

I do feel that you can test negative to Lyme and still have it. As you can see, it is possible for people to be diagnosed with Lyme disease while being persistently negative. It is also possible to be positive before treatment and have this decline to negativity later. And you can be positive and stay positive for decades. Patients who have longer active infection are more likely to be persistently positive.

So what does this mean in light of my previous comments on testing? The fact of the matter is that tests are forced to show a burden of proof in large scale trials before being validated by the FDA. They must be used in the manner described in order to be valid. The fact that you can test negative and have Lyme disease does not mean that western blots that show a few bands but not enough to be called positive are showing an active infection. Negative controls often show a few bands on western blots. It means better tests are needed.

But what about Lyme specialty labs? That’s getting its own post.



Akin E, McHugh GL, Flavell RA, Fikrig E, Steere AC. The immunoglobulin (IgG) antibody response to OspA and OspB correlates with severe and prolonged arthritis and the IgG response to P35 correlates with mild and brief arthritis. Infect Immun 1999;67:173-181.

Phillips SE, Mattman LH, Hulinska D, Moayad H. A proposal for the reliable culture of Borrelia burgdorferi from patients with chronic Lyme disease, even from those previously aggressively treated. Infection 1998;26:364-367

Marques AR, Stock F, Gill V. Evaluation of a new culture medium for Borrelia burgdorferi. J Clin Microbiol 2000;38:4239-4241

Tilton RC, Barden D, Sand M. Culture of Borrelia burgdorferi. J Clin Microbiol 2001;39:2747-2747

Bayer ME, Zhang L, Bayer MH. Borrelia burgdorferi DNA in the urine of treated patients with chronic Lyme disease symptoms: a PCR study of 97 cases. Infection 1996;24:347-353

Stricker RB, Johnson L. Persistent infection in chronic Lyme disease: does form matter? Research Journal of Infectious Diseases 2013.

Brian A. Fallon, Martina Pavlicova, Samantha W. Coffino, and Carl Brenner. A Comparison of Lyme Disease Serologic Test Results From 4 Laboratories in Patients With Persistent Symptoms After Antibiotic Treatment Comparison of Serologic Test Results. Clinical Infectious Diseases 2014: 59 (15 December) , 1705-1710.

Andrea T. Borchers, Carl L. Keen, Arthur C. Huntley, M. Eric Gershwin. Lyme disease: A rigorous review of diagnostic criteria and treatment. Journal of Autoimmunity 57 (2015) 82-115.

Fallon BA, Keilp JG, Corbera KM, Petkova E, Britton CB, Dwyer E, et al. A randomized, placebo-controlled trial of repeated IV antibiotic therapy for Lyme encephalopathy. Neurology 2008; 70:992-1003.

Krupp LB, Hyman LG, Grimson R, Coyle PK, Melville P, Ahnn S, et al. Study and treatment of post Lyme disease (STOP-LD): a randomized double masked clinical trial. Neurology 2003; 60:1923-30.

Marques AR, Stock F, Gill V. Evaluation of a new culture medium for Borrelia burgdorferi. J Clin Microbiol 2000; 38:4239-41.

Tilton RC, Barden D, Sand M. Culture of Borrelia burgdorferi. J Clin Microbiol 2001; 39:2747.

Bayer ME, Zhang L, Bayer MH. Borrelia burgdorferi DNA in the urine of treated patients with chronic Lyme disease symptoms: a PCR study of 97 cases. Infection 1996; 24:347-53.

Aguero-Rosenfeld, M. E., et al. Evolution of the Serologic Response to Borrelia burgdorferi in Treated Patients with Culture-Confirmed Erythema Migrans. Journal of Clinical Microbiology, Jan. 1996, p. 1–9.

Aguero-Rosenfeld, M. E., et al. Serodiagnosis in Early Lyme Disease. Journal of Clinical Microbiology, Dec. 1993, p. 3090-3095.

Glatz, Martin, et al. Clinical relevance of different IgG and IgM serum antibody responses to Borrelia burgdorferi after antibiotic therapy for erythema migrans. Arch Dermatol. 2006; 142(7):862-868.


Kalish, Robert A., et al. Persistence of Immunoglobulin M or Immunoglobulin G Antibody Responses to Borrelia burgdorferi 10-20 years after Active Lyme Disease. Clin Infect Dis. (2001) 33 (6): 780-785.


Aberer, E., et al. Course of antibody response in Lyme borreliosis patients before and after therapy. ISRN Immunology Volume 2012 (2012).

Lyme Disease: CDC Recommended Diagnostics (part 2)

Diagnosis of Lyme disease is a complicated affair. Part of the complication is the way our immune system responds to Lyme disease, and part of it is the current tests available to test for infection.

An antigen is anything that generates an immune response inside your body. Borrelia spp., the causative agents of Lyme disease, are antigens. They enter your body, your immune system recognizes it as a threat, and generates a specific IgM in response to Borrelia. This IgM is present in your serum about a week after infection, and persists in good quantities until about 12 weeks after infection, with concentration of specific IgM peaking around 4-6 weeks.

About 4 weeks after infection, your body generates Borrelia spp. IgG. This change from making IgM to IgG is called seroconversion. This IgG persists in good quantities for about 9-10 months after infection. While both IgM and IgG titers decrease over time, they can persist in serum for years after infection is resolved.

The standard Lyme diagnostics look for IgM and IgG. To be honest, I was pretty surprised at the abundance of different Lyme diagnostics, including others that look at that antigen and PCR based tests. There are multiple types of tests, and for each type of test, there are multiple companies that make them. It is a bit of a hot mess. So let’s look at the CDC recommended testing first, and then I’ll get to the others.

The CDC recommends a two step process for diagnosing Lyme. The first step is an ELISA and the second is a Western blot.

The Lyme ELISA (enzyme linked immunoassay) tests look for IgM and/or IgG, the antibodies made in response to the infection. These tests are sensitive, meaning that almost everyone with Lyme will test positive IF TESTED FOUR WEEKS AFTER SYMPTOMS PRESENT. (Note: I saw the phrase “almost everyone with Lyme will test positive” in relation to ELISA testing for Lyme several times. This test is most likely to correctly identify positives after patients have seroconverted.) However, these tests are not that specific, so they will provide 5-7% false positives. So with this test, if you have Lyme, you are likely to get a positive result, but if you don’t have it, you might still get a positive result. If the ELISA test is positive, then blot testing is done to confirm the result. If the ELISA test is negative, then the blot is not run, and the patient is told they do not have Lyme disease.

Western blots look for proteins specific to the organism in question. Western blots for Lyme testing are usually called immunoblots. These tests look for the specific IgG and IgM. Like the ELISA, the blot for IgG is more accurate, but requires the patient to have seroconverted. Blots for IgM are more likely to give false positives, so people who don’t have Lyme disease may have a positive test. The CDC guidelines recommend only running IgM blots in the first four weeks of illness due to the higher risk of false positive.

A Western blot works by showing “bands” to demonstrate that particular antigens were found. The bands are visible to the person reading the result. In order for an IgM blot to be considered positive, two of three possible bands must be positive. In order for an IgG blot to be considered positive, five of ten possible bands must be positive.


Let’s review.

You have had symptoms for two weeks. Your IgM ELISA is positive. Your IgG ELISA is negative (because your body hasn’t made IgG yet for Borrelia spp.) Your IgM blot shows 3/3 bands. Your IgG blot is negative. You are diagnosed with Lyme disease.

You have had symptoms for two weeks. Your IgM ELISA is positive. Your IgG ELISA is negative (because your body hasn’t made IgG yet for Borrelia spp.) Your IgM blot shows 1/3 bands. Your IgG blot is negative. You are not diagnosed with Lyme disease.

You have had symptoms for two weeks. Your IgM ELISA is negative. Your IgG ELISA is negative (because your body hasn’t made IgG yet for Borrelia spp.) No blots are run. You are not diagnosed with Lyme disease.

You have had symptoms for six weeks. Your IgM ELISA is positive. Your IgG ELISA is positive. Your IgM blot shows 2/3 bands. Your IgG blot shows 5/10 bands. You are diagnosed with Lyme disease.

You have had symptoms for six weeks. Your IgM ELISA is negative. Your IgG ELISA is positive. Your IgM blot is negative. Your IgG blot shows 5/10 bands. You are diagnosed with Lyme disease.

You have had symptoms for six weeks. Your IgM ELISA is negative. Your IgG ELISA is positive. Your IgM blot is positive. Your IgG blot shows 4/10 bands. You are not diagnosed with Lyme disease.

You have had symptoms for six weeks. Your IgM ELISA is positive. Your IgG ELISA is negative. Your IgM blot shows 3/3 bands. Your IgG blot shows 4/10 bands. You are not diagnosed with Lyme disease.


So let’s talk about this two step process. It has been studied, a lot. The two step process was implemented in the mid-90’s due to the lack of specificity, meaning that people were being treated for Lyme disease who didn’t have it. When the two step process is run and interpreted by skilled operators, according to recommendations, it has 99% specificity. This means a 1% chance of calling someone negative when they really have Lyme disease.

But this doesn’t seem to translate well to clinical practice. A key weakness of these diagnostics in practice is their inability to correctly diagnose people who are newly infected. One study found that the sensitivity and specificity for the two tier test was 100% and 99% for people who no longer had the bull’s eye rash, which happens during the acute phase. But it only had 29% sensitivity for patients in the acute phase with the rash. (Steere 2008) Another study looking particularly at clinical practice found that 50/182 patients had a false positive IgM blot. 78% of them received unnecessary antibiotics. (Seriburi 2012)

I understand why these tests work fine in some studies and not in others. As far as ELISAs giving false positives, that’s no surprise to anyone who has run them before. They’re “sticky.” They crossreact with a lot of things. The reagents are optimized to minimize this, but it is a persistent problem with this type of test. Instead of getting an exact match for your target (Borrelia spp.), you get a “close enough.” You get a positive result if you get a close enough match, and things that match close enough include antibodies for anaplasmosis, leptospirosis, syphilis, Epstein Barr virus, and lupus, among others.

Western blots are also problematic. They are time consuming and complicated. There are a lot of steps and there is a lot of repetition. Reading the results can be confusing. You are comparing the intensity of a band to a control. So let’s say the darker it is, the more positive it is. Just seeing a band does not make it positive. It has to also be as dark as the control for the same band. And things like darkness are subjective.

A sticking point for a lot of people seems to be the fact that they have some bands on their Western blots but not enough to meet the positive criteria. A lot of people feel that if they are positive for some bands that this should be sufficient for a diagnosis. I understand why this is confusing. It’s confusing because it seems like if these blots test for proteins specific to Lyme disease that having any of them should be indicative of infection. But that’s not true.

Remember how I said ELISA tests are sticky, that you can get a good enough match that’s not specific? There’s some of that happening in Western blotting, too. And remember how I said that the procedure is difficult, that the results are subjective? Those facts mean that you can get a few bands through operator error. That’s why it is so important that ALL the required bands show up.

Diagnostic development involves employing a lot of measures to ensure you compensate for errors borne out of biologic crossreaction (like sticky antibodies) and procedural inadequacies. The result is very specific criteria for what is considered positive and what is not. The FDA requires this.

Another thing to keep in mind for later is that diagnostics that are FDA validated are designed for very particular uses. That means that if you have an FDA validated test for Lyme IgG in serum and you use that test to test whole blood or cerebrospinal fluid or synovial fluid, and you get a positive result, it is meaningless. It is not positive. It is nothing. If you have an FDA validated test that tells you to use 100 ul of serum and 100 ul of something else per tube, and you use 150 ul of serum and 150 ul of something else per tube, and you get a positive result, it is meaningless. This also generates a lot of confusion because people feel like if you are looking for the same thing (Lyme IgG), it shouldn’t matter where you find it. But it does.

It matters because sample matrices (like serum or whole blood or whatever) have their own specific “backgrounds.” This means that they contain different proteins and have different fluid dynamics and they could cause additional crossreaction or they could cause less crossreaction. The chemistry involved in diagnostics is very fine. Tests that have a large tolerance for changes like this are called “robust.” The Lyme diagnostics discussed here are not robust.



Steere AC, McHugh G, Damle N, Sikand VK. Prospective study of serologic tests for Lyme disease. Clin Infect Dis. 2008;47(2):188–195. doi: 10.1086/589242.

Wormser GP, Carbonaro C, Miller S, Nowakowski J, Nadelman RB, Sivak S, Aguero-Rosenfeld ME. A limitation of 2-stage serological testing for Lyme disease: enzyme immunoassay and immunoblot assay are not independent tests. Clin Infect Dis. 2000;30(3):545–548. doi: 10.1086/313688

Klempner, M. S., C. H. Schmid, L. Hu, A. C. Steere, G. Johnson, B. McCloud, R. Noring, and A. Weinstein. 2001. Intralaboratory reliability of serologic and urine testing for Lyme disease. Am. J. Med. 110:217-219

Reed, Kurt D. Laboratory Testing for Lyme Disease: Possibilities and Practicalities. J Clin Microbiol. Feb 2002; 40(2): 319–324.

Ang, C. W., et al. Large differences between test strategies for the detection of anti-Borrelia antibodies are revealed by comparing eight ELISAs and five immunoblots. Eur J Clin Microbiol Infect Dis. Aug 2011; 30(8): 1027–1032.

Woods, Charles R. False-Positive Results for Immunoglobulin M Serologic Results: Explanations and Examples. J Ped Infect Dis (2013).

Seriburi, N. Ndukwe, Z. Chang, M. E. Cox and G. P. Wormser . High frequency of false positive IgM immunoblots for Borrelia burgdorferi in Clinical Practice. Clin Microbiol Infect 2012; 18: 1236–1240

Branda, John A, et al. 2-Tiered Antibody Testing for Early and Late Lyme Disease Using Only an Immunoglobulin G Blot with the Addition of a VlsE Band as the Second-Tier Test. Clin Infect Dis. (2010) 50 (1): 20-26.