How Is Acute Porphyria Diagnosed?
The Porphyria's are solely diagnosed by Doctor, Nurse Practitioner, or Physicians Assistant, based on laboratory testing, and the patients signs and symptoms. The Acute porphyria's consists of testing porphyrin's in blood, stool, and urine (Janus MD PhD, 2017) (with porphyrins and porphyrin precursors ALA and PBG), and the enzymes associated with each porphyria (certain enzyme testing is only currently available at the research level in the US).
Porphyrins are chemicals that help make hemoglobin, a type of protein in your red blood cells. Hemoglobin carries oxygen from your lungs to the rest of your body.
*These are not standard lab tests, we wince when people say "all my tests were normal", the truth is all routinely done tests were (close to) normal, these aren't routinely done tests. Our organization is in agreement with The Australian Porphyria expert Dr. Janus who advises to test each patients blood, (plasma), urine, and stool. And an American porphyria expert weighs in: "Measurement of enzyme activities... are important for diagnosis". (Anderson, Acute Hepatic Porphyria current diagnosis and management, 2019)
The "stepwise approach" (recommended elsewhere) is incredibly dangerous as it forces patients to wait for an attack, which is unnecessary, and tests only urine initially, which will miss younger patients, many latent patients, and may miss (a high percentage of) HCP, and VP, in life threatening attacks, according to Dr. Janus and a flurry of other experts "for some PBG and ALA in urine levels may be normal during and following an attack especially in HCP" (Janus, 2017).
In "Family Studies of AIP and HCP in Nikata and Akita Prefectures, Japan" Dr. Sasaki notes: "the urinary PBG test was negative in all 108 members of HCP families."
Blake's "Fecal Coproporphyrin Isomers in Hereditary Coproporphyria" found over 30% of HCP patients in his study didn't have even high fecal porphyrins (much less elevated urinary porphyrins), and in the drug trial for Panhematin it was noted the VP patient had a false negative urine PBG test in an attack (Peterson, 1976 "Hematin for treatment of Acute Porphyria"), in an early (Pan)hematin drug trial.
Please see cautions below, there is a high rate of false negatives (Official guideline for the Porphyria Association Inc, By Professor Janus updated 2017).
1.) Blood test of the Enzyme Porphobilinogen Deaminase (Aminolevulinic acid Dehydratase may also be done in the US).
The following tests must be protected from light and heat:
2.) Fecal Porphyrins, fractionated (test must be High Performance Liquid Chromatography)
3.) Porphyrins Plasma fluorescence scan
4 & 5.) Urine Aminolevulinic acid (ALA) and Porphobilinogen (PBG)
6.) Total urine porphyrins fractionated and quantified
Does not have to be heat and light protected:
7.) DNA testing
We give cautions and tips below, porphyria is a metabolic disease and false negatives often occur.
1. Enzyme activity
There are three enzyme activity tests available in the US, two are for the Acute Porphyria's:
The enzyme test for AIP called Porphobilinogen Deaminase "PBGD" was found to be as effective identifying 84% of patients with AIP as testing the Urine for elevated Porphobilnogen "PBG" which identified 85% of AIP patients in remission (Kaupinnen, 2002 "Molecular and Biochemical studies of AIP in 196 Patients and their families") . The positive about the enzyme test PBGD is it can be done anytime without endangering the patients life by forcing them to wait for an attack. Low PBGD may also occur in patients with VP (Doss, 2001 "VP with coexistent decrease in PBGD activity") and in patients with HCP or VP (Anderson, 2019 "Safety and Efficacy of Panhematin...").
Acute Porphyria Enzyme tests (available commercially in the US)
1. Porphobilinogen Deaminase (PBGD) is AIP's enzyme, but can be low in active HCP and VP
2. Aminolevulinic acid dehydrastase (ALAD) ALAD- Porphyria.
Chronic Hepatic Cutaneous Porphyria Enzyme test:
3. Uroporphyrinogen decarboxylase (UROD) PCT's enzyme
a. In 5-16% of AIP patients the PBGD enzyme is not low due to alternate DNA splicing (Kaupinnen 2002).
b. In symptomatic/Active AIP patients the enzyme PBGD can raise to reference/ normal levels.
c. Other illnesses may cause a low PBGD such as sideropenia, and the following illnesses may affect PBGD levels: anemia (this is often seen in Porphyria), uremia, malignancies, and hemodialysis ( Kaupinnen 2002.".AIP in 196 patients.."
d. During an attack the level of PBGD can rise to reference levels creating a false negative (Kostreswa 1986 "Increased activity of PBGD in erythrocytes during attacks in AIP".
e. Below normal PBGD is found in some active HCP and VP patients (Anderson, Efficacy of Panhematin, 2019).
f. Both the PBGD test and the Aminolevulinic Acid dehydratase Enzyme Whole Blood
must be refrigerated and best done outside of an attack.
g. While ALAD Porphyria is very rare, about 1-2% of the population have below normal ALAD enzyme, which might be caused by another acute porphyria (or some other cause), similar to the reduction in PBGD in HCP and VP, so please request a through work up if you have low ALAD enzyme, and take any signs and symptoms consistent with an attack seriously.
h. Low UROD may not be seen in all PCT.
i. There are multiple enzyme activity tests missing in the US which are offered in other countries that our organization would like to offer, they are HCP's enzyme coproporphyrinogen oxidase (CPOX), and VP's enzyme protoporphyrinogen oxidase (PPOX), uroporphyrinogen synthase (UROS), ferrochelatase (FECH). We are working to bring those tests here, join us!
2. Fractionated fecal porphyrins
Stool tests for porphyria includes total fecal porphyrins (quantity) and fractionated fecal porphyrins (quality).
a. It is important in to make sure both total fecal porphyrins (quantity) and Fractionated fecal porphyrin (quality/ ratio) are tested.
b.The sample must be light protected (container wrapped in foil, or black container) immediately, and frozen or refrigerated per labs requirements.
c. Children may have false negative results below the age of 12 (Blake, 1991) and by age 10 a CIII:CI ratio was detectable (Allen, HCP comparison of molecular and biochemical investigations in large family).
d. Fecal Coproporphyrin dominance (CIII+CI) is an ABNORMAL finding and is only seen in one Acute Porphyria, Hereditary Coproporphyria (Singal, 2013 "Variegate Porphyria"), and is also not seen in normal "non porphyric" humans (Doss, 1999 "Compound heterozygous HCP"). In a normal fecal porphyrins profile the coproporphyrin (CIII+CI) is 25% of total fecal porphyrins (Australasian pathology).
e. Patients with HCP may not have high porphyrin excretions in fecal testing (Blake, 1991 "Fecal Coproporphyrin Isomers in Hereditary Coproporphyria) 40% of one family with HCP did not have high fecal porphyrin excretion.
f. Even without a CIII:CI inversion (seen in many HCP) a coproporphyrin dominant fecal sample is abnormal, not found in normal non porphyric humans (Doss, 1999 and Australasian Pathology) and coproporphyrin dominance is only seen in HCP (Singal, 2013).
g. The fecal porphyrins test can also detect VP in some cases, which may be elevated, and is mostly Protoporphyrin IX (UTMB, Porphyria lab) .
h. A fairly large fecal sample is needed 10 grams is recommended by one lab.
*Unfortunately US labs are not currently flagging some abnormal results, if you see that your results are abnormal, but not flagged, we can help you report the lab to the proper governing body, which is CLIA, or the College of American Pathologist, please reach out to us.
3. Plasma tests for porphyria consists of total plasma porphyrins, fractionated plasma porphyrins, plasma fluorescence, erythrocyte porphyrins.
The total plasma porphyrins
test identifies the totality of porphyrins present in plasma (citation). The most common method for measuring total plasma porphyrins is (citation).
The fractionated plasma porphyrins test separates the totality of porphyrins present in plasma into individual components (citation). The most common method for separating porphyrins in plasma is high-performance liquid chromatography (HPLC), which is a laboratory technique used to separate, identify, and quantify each component in a mixture.
a. the test is light sensitive, the test tube must be covered by foil or dark paper during the blood draw.
b. Maybe positive in symptomatic HCP (30%) and AIP, if the patient has VP, is old enough- age 22+, and has had an attack, a majority- but not all VP, will show a 625, ( University Cape Town Porphyria) 625, or 626, or 627 wave length.
c. 80 % of VP patients will have a positive plasma scan (University of Cape Town Porphyria).
4, 5, & 6 Urine tests for porphyria includes total urine porphyrins and urine precursors (ALA and PBG) and fractionated urine porphyrins.
Urine precursors (ALA and PBG)
Urine Aminolevulinic acid (ALA), urine Porphobilinogen (PBG), urine total porphyrins, urine fractionated porphyrins ,
*ALA / PBG Urine
Aminolevulinic acid is the best test for ALAD-P, and is also used in conjunction with PBG testing for AIP,
(16 listed below)
a. If testing urine please test the first morning urine of the day, ("How to test for Porphyria" Official guidelines for the Porphyria association Dr Janus MD PhD) or 24 hr. test, when symptomatic.
b. All samples must be protected from light; urinary porphyrin concentrations can decrease by up to 50% if kept in the light for 24 hours (Deacon et al. 2001). Keep the sample refrigerated or frozen per labs guidelines.
c. A concerning number of HCP and VP patients are well documented in research papers to never have a positive urine PBG test in and out of an attack, it is simply incomplete and dangerous to do only urine testing, and also possibly illegal in the US, the Urine PBG test is not FDA approved test, and may not be used as a sole means of diagnosis. An Australian researcher Dr Janus, found that in HCP neither the urine ALA or PBG may be elevated in an attack, and VP patients self report the urine testing is often inconsistent and false negative with attacks.
d. Younger people with any type of Acute Porphyria, symptomatic or not, and HCP, and VP (regardless of age) may not have positive urine ALA or PBG tests even in attacks (Janus). The French Porphyria Center states in AIP false negatives occur ages 15 and lower. The South Africa Uni. of Cape Town found in VP under the age of 22 may have false negative testing VP (the problem of silent carriers) and VP patients themselves report the urine testing is inconsistent with their attacks.
e. A fair percentage of AIP patients can have elevated PBG / and or ALA outside of attacks. Kaupinnen found that intra-individual increases in attacks was about 1.6-4 (Kaupinnen 2002 "..196 patients with AIP"). Anderson states that attacks in VP and HCP are clinically diagnosed (Anderson 2019 Efficacy..Panhematin study) It is extremely important to go by what the patient is telling the medical professional, the testing doesn't always reflect the attack.
f. Children can have very serious acute porphyria attacks, and they are documented to occur with negative PBG urine testing. (Singh, 2003 AIP: An unusual case of hypertension.)
g. Once a patient has a clear porphyria diagnosis there is no need to continue testing ("How to test for Porphyria" Official guidelines for the Porphyria association Dr. Janus MD PhD updated 2017). Patients have shared horror stories how they had a positive test PBG urine test in an attack and improved with treatment, and then lost their treatment because a further test was a false negative. Attacks are clinically diagnosed. (Anderson, 2019)
h. Certain antibiotics (methenamine hippurate) can cause false negatives on the urine PBG test (European Porphyria Network).
i. I.V. Glucose/ carbohydrates may cause a false negative urinary PBG test (Doheny, 2016)
j. A patient on "Porphyria Sucks" a face book support group in 2018 shared that she has HCP, and the genetics laboratory found she also has a porphyrin retention mutation (2018 Porphyria Sucks face book Support Group)
k. For HCP and VP, urine PBG and ALA levels may be less elevated during attacks than in AIP and are more likely normal between acute episodes (Balwani"AHP recommendation for evaluation.." ).
l. A very dilute urine sample that may lead to ‘false-negative’ spot urine if not normalized to urine creatinine (Balwani"AHP recommendation for evaluation.." ).
m. Plasma PBG and ALA testing was found to be a better test than the urine ALA/ PBG test in HCP and VP (the research paper is cited elsewhere in this website).
n. Patients in renal failure can be plasma tested for PBG and ALA instead of urine. (University of Texas Medical Branch Porphyria Lab).
o. Because urine may contain substances that inhibit or mimic the PBG-Ehrlich aldehyde reaction, quantification of PBG requires prior purification by anion-exchange chromatography.". (Elder, Investigation of the Porphyria's)
7. Genetic testing
can be done with Saliva or blood
The DNA tests that are available are: next-generation sequencing, sanger sequencing, and microarray-based comparative genomic hybridization. All genetic testing in the US requires a Doctors signature or "order".
The Acute Porphyria genes are: ALAD, CPOX, HMBS, PPOX,
The other porphyria genes are: ALAS2, C15orf41, FECH, HFE, SLC19A2, UROD, UROS.
a.) Unlike the Breast cancer gene (BRAC1) which in found in large groups unrelated people, most porphyria mutations are "family specific" (Singal, "Variegate Porphyria" 2013) also called "single family mutations" (Capellini 2001 "Hematologically important mutations Acute Intermittent Porphyria"), found in only one or two families, except for the S. African variant, and a few variants in N. Europe, which are considered founder effect variants and are shared among large numbers of people. There is no common American or N. American Acute Porphyria variant. A patient can have a negative DNA test and have porphyria, this happens in 1 of 5 patients, with just 1 type of porphyria test, according to one porphyria genetics counselor (Doheny, 2016) , this is probably a much higher number, given there are 4 other tests, and only the urine test was used.
b. Genetic testing is relatively new and not 100% reliable, if your families mutation has not been found, you may have porphyria, but have a negative genetics test. A negative genetics test does not clear you of porphyria,
c. 13% of the patients had a positive genetics test without any positive biochemical testing (Doheny, 2016).
d. A negative genetics test- because they are not FDA approved, may not be used to deny treatment, or strip a diagnosis, it just means if you have porphyria that your variant has not been discovered yet.
e. It is important to note that labs that do not offer deletion and insertion testing might not find mutations with large deletions or insertions. At this time it is not clear how often deletion and insertion mutations occur. One source says that approximately forty percent of genetic mutations for Hereditary Coproporphyria are deletions or insertions, whereas another source says that approximately five percent of genetic mutations for Acute Intermittent Porphyria are deletions or insertions.
f. Genetics testing may only catch 20% of gene carriers (Rehm, H. PhD, Harvard 2018) .
g. We encourage all to request your underlying DNA mutations, as there are variants of "uncertain significance" under HIPPA regulations you are legally entitled to receive those within 30-60 days of written request to the laboratory.
h. At least one AIP variant is found on a different gene than the HMBS gene.
There are a limited number of laboratories in America that test for porphyria, so you or your doctor will have to send the samples to a laboratory or multiple laboratories that test for porphyria across the country. We want to make it very clear no one test is 100% accurate, and certain porphyria tests have false negative rates of close to 40%.
Porphyria is an inborn error of metabolism and as such testing has to be compared to clinical history. As Acute Porphyria is a life threatening disease we encourage a full battery of testing which may have to be repeated until you the patient are satisfied. The current average time to an Acute Porphyria diagnosis is 15 years in the U.S.,our organization would like to shorten that to days.