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Inflammatory Process
Blood Amino Acids
Blood Fatty Acids
Blood Nutrient/Toxic Elements
Blood Food Antibody   
Blood ADMA
Blood Vitamin D
Blood Lymphocyte Growth Assays
Blood Lipid  
Blood Antioxidant Capacity
Blood Oxidant Stress Markers (e.g., Glutathione)
Blood Hs-CRP, Ferritin, and/or Fibrinogen
Blood SNP Genomic Tests
Urinary Toxic Metals (provoked and unprovoked)
Urinary Organic Acids
Urinary Oxidant Stress Markers
Fecal Lysozyme
Fecal Lactoferrin
Fecal Calprotectin

Hormone and Neurotransmitter Regulation
Blood Amino AcidsBlood Fatty Acids
Blood Food Antibody
Blood Estrogen, Progesterone, Testosterone, DHEA, SHBG
Blood Estrogen Metabolites
Blood Vitamin D
Blood Lymphocyte Growth Assays
Blood Lipid
Blood Thyroid Function (T3, T4, TSH, RT3, antibody)
Blood Insulin/Glucose
Blood SNP Genomic Tests
Urinary Organic Acids
Urinary Toxic Metals (provoked and unprovoked)
Urinary Amino Acids
Urinary Estrogen Metabolites
Urinary Iodide
Urinary Neurotransmitter Metabolites
Salivary Estrogen, Progesterone, Testosterone
Salivary Cortisol, DHEA
Salivary Melatonin

Digestion, Absorption and Barrier Integrity  
Blood Fatty Acids
Blood Food Antibody
Blood H. Pylori
Blood Candida
Antibody/Antigen
Blood Celiac Profile
Blood Lymphocyte Growth Assays
Blood Lipid  
Urinary Organic Acids
Urinary Lactulose/Mannitol
Fecal Lysozyme
Fecal Lactoferrin
Fecal Ova & Parasites
Fecal Bacteriology Culture
Fecal Short-Chain Fatty Acids
Fecal Digestive Elements
Fecal Absorptive Elements
Fecal Mycology Culture
Fecal H. Pylori Antigen
Fecal Clostridium Difficile Toxins A & B
Fecal Enterohemorrhagic E. Coli Cytotoxin 
Fecal Elastase-1
Fecal Calprotectin
Breath Lactose Intolerance
Breath Small Intestine Bacterial Overgrowth

Detoxification and Biotransformation
Blood Amino Acids
Blood Nutrient/Toxic Elements 
Blood Estrogen Metabolites
Blood Oxidant Stress Markers (e.g., Glutathione)
Blood Lymphocyte Growth Assays
Blood Antioxidant Capacity
Blood SNP Genomic Tests
Urinary Organic Acids 
Urinary Amino Acids
Urinary Toxic Metals (provoked and unprovoked)
Urinary Estrogen Metabolites
Urinary Challenge Detox Profiles
Urinary Mercapturic and D-Glucaric Acids
Urinary Oxidant Stress Markers
Fecal Toxic Metals
Hair Toxic Metals

Oxidative/Reductive Homeodynamics
Blood Amino Acids     
Blood Nutrient/Toxic Elements
Blood ADMA
Blood Estrogen Metabolites
Blood Vitamin D
Blood Oxidant Stress Markers (Glutathione)
Blood Antioxidant Capacity
Blood Lymphocyte Growth Assays
Blood SNP Genomic Tests
Urinary Organic Acids
Urinary Oxidant Stress Markers
Urinary Amino Acids
Urinary Toxic Metals (provoked and unprovoked)
Urinary Estrogen Metabolites
Urinary Challenge Detox Profiles

Immune Surveillance
Blood Amino Acids     
Blood Nutrient/Toxic Elements
Blood Food Antibody
Blood Vitamin D
Blood Mold/Yeast Antibody
Blood Celiac Profile
Blood Lymphocyte Growth Assays
Blood Antioxidant Capacity
Blood SNP Genomic Tests
Urinary Organic Acids
Urinary Amino Acids
Urinary Toxic Metals (provoked and unprovoked)
Fecal Secretory IgA

Psychological and Spiritual Equilibrium
Blood Amino Acids     
Blood Nutrient/Toxic Elements
Blood Lymphocyte Growth Assays
Blood Antioxidant Capacity
Blood Fatty Acids
Urinary Organic Acids
Urinary Amino Acids
Urinary Toxic Metals (provoked and unprovoked)
Hair Toxic Metals

Structural Integrity
Blood Nutrient/Toxic Elements
Blood Vitamin D
Blood Lymphocyte Growth Assays
Blood Antioxidant Capacity
Blood Lipid
Urinary Organic Acids
Urinary Toxic Metals (provoked and unprovoked)
Urinary Bone Resorption
Urinary Amino Acids

Part II: Brief Descriptions of Functional Laboratory Tests

(Tests are organized alphabetically using the names shown in Part I above.)

Blood ADMA

Plasma asymmetric dimethylarginine (ADMA) is a primary endogenous inhibitor of nitric oxide synthase that inhibits nitric oxide production. Elevated levels are implicated in cardiovascular disease and are a cause of hypertension, hyperhomocysteinemia, diabetes mellitus types 1 & 2, and pre-eclampsia. ADMA becomes elevated due to loss of activity for the ADMA-degrading enzyme DDAH when antioxidant protection is lacking.

Blood Amino Acids

Amino acid analyses aid in the assessment of: dietary protein adequacy and balance, gastrointestinal dysfunctions, forms of protein intolerance, nutritional deficiencies (vitamins, minerals), renal and hepatic dysfunction, psychiatric abnormalities, susceptibility to inflammatory response and oxidative stress, reduced detoxification capacity, susceptibility to occlusive arterial disease, and many inherent disorders of amino acid metabolism. Many free amino acids such as histidine, lysine, threonine and serine in plasma provide toxic element ligands for transport and renal excretion. Amino acid hepatic flux stimulates carcinogenic toxicant conjugation reactions. Amino acid analysis also shows phase II biotransformation substrates (glycine and sulfur amino acids) and status of glutathione precursor amino acids methionine and glycine. Fasting plasma and whole blood (bloodspot) levels provide a reliable assessment of dietary adequacy. Plasma hydroxylysine and hydroxyproline become elevated due to increased rates of bone resorption. 

Blood Antioxidant Capacity

T-lymphocytes are cultured under optimal growth conditions and, following mitogenic stimulation, are exposed to increasing concentrations of free radicals to measure the cell’s ability to resist oxidative stress.  Specific antioxidants (i.e., selenium, tocopherols, glutathione, Coq10, etc.) are available to identify specific intracellular functional deficiencies that may limit antioxidant capacity within a highly interactive system of antioxidants.

Blood Candida Antibody/Antigen

Serum IgM, IgG, IgA antibodies for Candida albicans are used to indirectly assess current, past, and mucosal exposure to this yeast, respectively. Serum Candida albicans (antigen) is measured to identify its presence in circulation. Candida is associated with both acute and chronic conditions.

Blood Celiac Profile

Total immunoglobulins A, tissue transglutaminase IgA, and anti-gliadin IgA are measured. An elevated tissue transglutaminase IgA is highly sensitive and specific for gastrointestinal villous atrophy associated with Celiac. An elevated anti-gliadin IgA indicates specific gluten-activated immune response.

Blood Estrogen Metabolites

Blood estrogen metabolites include 2-hydroxyestrone and 16-alpha-hydroxyestrone, and reflect how estrogen is being processed in the body. Blood sampling provides a direct assessment of circulating estrogen metabolites that act directly on target tissues. Imbalances of these metabolites are associated with cardiovascular disease, depression, osteoporosis, lupus, and breast cancer.

Blood Estrogen, Progesterone, Testosterone, DHEA, SHBG

Serum assays provide an accurate and time-tested assessment of estrogen, progesterone, testosterone, and DHEA at a point in time. Sex hormone binding globulin (SHBG) is also measured in serum to provide an estimate of free testosterone and estradiol, which are primarily bound by this protein. These hormones are assessed to evaluate ovarian and adrenal dysfunction, and to determine need and dose of related hormone replacement therapy. 

Blood Fatty Acids (plasma, RBC, blood spot)

Fatty acid (FA) profiles show quantitative results for individual fatty acids, including polyunsaturated omega-3 (including ALA, EPA, DHA), omega-6 (including GLA, DGLA, AA), omega-9 (including oleic, nervonic), saturates (including capric, palmitic, stearic, hexacosanoic), and trans-fatty acids. Fatty acid ratios such as LA/DGLA and AA/EPA are calculated to help identify imbalances. Essential FA deficiencies and metabolic effects manifest as imbalances within and between families of fatty acids.

 

Blood Food Antibody

Semi-quantitative levels of serum IgE or IgG subclass responses to 30-90 common foods are used to identify food allergy or sensitivity due to leaky gut and to customize elimination/rotation diets.

Blood H. pylori

Helicobacter pylori is the bacterium that causes peptic ulcer disease and is associated with increased risk of gastric cancer.  Blood assays indirectly identify the presence of H. pylori by measuring antibody levels in circulation. 

Blood Hs-CRP, Ferritin, Fibrinogen

Testing for these acute phase protein markers in serum provides insight relevant to the inflammatory cascade, free iron modulation, and the clotting cascade, respectively.

Blood Insulin/Glucose

Elevated fasting serum insulin indicates insulin insensitivity that produces increased serum triglycerides and LDL cholesterol. Euglycemia in this scenario is characteristic of the metabolic syndrome, which is associated with increased risk of cardiovascular disease and diabetes

Blood Lipids 

Cardiovascular disease is a multifactorial process whose etiology encompasses both size and number of various lipoproteins. These include HDL and LDL subclasses, remnant lipoprotein, VLDL, and lipoprotein (a).   Homocysteine, although not a lipoprotein, is recognized as a contributing factor to hemostasis, neurological function, and cardiovascular disease, and is easily measured in blood lipid studies. 

Blood Lymphocyte Growth Assays

T-lymphocytes are grown in tissue culture by mitogenic stimulation of proliferation.  By culturing the cells under a variety of conditions, functional deficiencies of vitamins, minerals, amino acids, antioxidants and other micronutrients can be assessed.  The assays are a long-term indicator of nutritional status and are useful not only in the identification of functional deficiency states, but also in monitoring the effectiveness of repletion to achieve optimal intracellular function of the cofactors within cellular metabolism.

Blood Mold and Yeast 

Mold and yeast blood antibody testing is used to assess for the immunologic reaction to these potential toxins. Mold and yeast exposures are associated with various acute and chronic conditions. 

Blood Nutrient and/or Toxic Elements

Whole blood element analysis is a diagnostic method that assists in determining deficiencies, excesses, and imbalances of essential elements, as well as recent or ongoing exposure to specific toxic elements. Whole blood analysis measures total element levels that circulate extracellularly (serum/plasma) as well as intracellularly (function within blood cells). RBC element levels are very useful for assessing: cardiotonic influences (magnesium, potassium); anti-inflammatory processes (selenium, copper, zinc); anemia (copper, iron); immunological function (zinc, copper, magnesium); and glucose tolerance (chromium, manganese, and possibly vanadium). Accurate assessment of essential element status aids in determination of appropriate supplementation.

Blood Oxidant Stress Markers

Plasma amino acid testing reveals ability to maintain fasting blood levels of glutathione precursors methionine, homocysteine and glycine (taurine adds a further assessment of sulfur metabolism). Total body oxidant stress is evaluated with serum lipid peroxides. Erythrocyte elemental analysis is used to assess essential elemental cofactors, selenium, zinc, copper and manganese for redox-balancing enzymes like superoxide dismutase, and elevation of toxic elements that contribute to oxidant stress.

Blood SNP Genomic Tests

Genomic testing identifies specific variations in nucleic acid sequencing called single nucleotide polymorphisms (SNPs). SNPs can influence many different functions:

Detoxification: SNPs can alter the kinetics of enzyme activity, affecting phase I and phase II detoxification and folic acid activation. Identification of these SNPs may indicate the need for lifestyle adjustments and nutritional supplementation that support normal enzyme activity, and highlight environmental exposures that possess increased toxicity.

Hormone and neurotransmitters: SNPs that affect induction and activity of enzymes that metabolize hormones and neurotransmitters may increase risk for related cancers and mood disorders. Identification of these SNPs may indicate the need for lifestyle adjustments and nutritional supplementation that support normal enzyme activity.

Immune: SNPs that affect the levels and activity of cytokines can modulate immune and inflammatory activity. These variations can affect balance between cell (Th-1) and humoral (Th-2) immunity and reveal potential defects in immune system defense.

Inflammatory: SNPs that affect the levels and activity of cytokines can modulate immune and inflammatory activity. These variations can affect balance between cell (Th-1) and humoral (Th-2) immunity, reveal potential defects in immune system defense, and stimulate mechanisms leading to chronic, overactive inflammatory responses. Identification of these SNPs may indicate the need for lifestyle adjustments and nutritional supplementation that support normal cytokine activity. 

Oxidative stress: SNPs that affect the production or management of oxidative stress are associated with chronic illness. SNPs can alter protein induction and activities that elevate oxidative stress, including phase I and II detoxification enzymes, superoxide dismutase, and cytokines.

Blood Thyroid function (T3, T4, TSH, RT3, and antibody)

A comprehensive thyroid function analysis includes free T3, free T4, TSH, reverse T3, as well as thyroid Peroxidase and thyroglobulin antibodies. These tests are used to diagnose thyroid disease, and to determine need for thyroid support, including need and dose of related thyroid hormone therapy. 

Blood Vitamin D

Serum 25-hydroxyvitamin D is the major circulating precursor to active 1, 25-dihydroxy vitamin D. Low values show vitamin D deficiency and repeat testing allows monitoring of repletion in patients with osteoporosis or chronic inflammatory disorders.

Blood Vitamin Profile

Measurement of serum concentrations of fat-soluble vitamins including A, E, CoQ10, beta-carotene, and lycopene allow direct assessment of dietary sufficiency for these key antioxidants, essential or conditionally essential nutrients.

Breath Lactose Intolerance

This breath test identifies bacterial fermentation of lactose in the GI and indicates presence and severity of lactose intolerance.

Breath Small Intestine Bacterial Overgrowth

This breath test identifies overgrowth of bacteria in the small intestine. 

Fecal Absorptive Elements

Long-chain fatty acids, phospholipids, cholesterol, triglycerides, and total fecal fat are measured in the stool to assess malabsorption. When poorly absorbed, these fecal fats will be elevated in the stool. 

Fecal Bacteriology Culture 

Bacteria are cultured from stool samples to identify the presence and relative abundance of specific bacteria, both beneficial and dysbiotic. When dysbiotic bacteria are present, further testing is performed to determine which natural and pharmaceutical agents might eradicate these specific bacteria.

Fecal Calprotectin

Calprotectin is a neutrophil-derived protein that is released in the GI tract in the presence of infectious, inflammatory, or malignant disease. This quantitative assay can identify mild, moderate and severe levels of inflammation, and is FDA approved to differentiate irritable bowel syndrome (IBS) from inflammatory bowel disease (IBD). It can also be used to assess the effectiveness of treatment and to predict relapse in IBD cases.  

Fecal Clostridium Difficile Toxins A & B

The C. difficile toxin enzyme immunoassay test is the standard diagnostic tool for identification of overgrowth of C. difficile bacteria in the gastrointestinal tract.

 

Fecal Digestive Elements

Pancreatic elastase 1, chymotrypsin, short-chain fatty acids, meat fibers, and vegetable fibers are used as an indirect evaluation of digestive function. These markers provide insight into inadequate digestive enzyme production and maldigestion. 

Fecal Elastase

Elastase, produced in the pancreas, is a proteolytic enzyme that breaks down dietary protein to absorbable amino acids and peptides. Fecal elastase levels are utilized to diagnose poor digestive capacity due to pancreatic exocrine function that may be associated with chronic pancreatitis, cystic fibrosis, carcinoma of the pancreas, type 1 diabetes mellitus, and other etiologies of pancreatic enzyme insufficiency.

Fecal Enterohemorrhagic E. coli Cytotoxin

Measurement of these specific E. coli cytotoxins in the feces is a reliable tool for the diagnosis of EHEC infections.

Fecal H. pylori Antigen

The fecal HpSA enzyme immunoassay (EIA) is a qualitative procedure for the detection of H. pylori antigens in stool. This noninvasive test’s specificity and sensitivity for diagnosis of H. pylori infection are comparable to blood and tissue biopsy methods. The fecal H. pylori antigen is also excellent for assessing eradication treatment efficacy. 

Fecal Lactoferrin

A highly sensitive and specific immunoassay has recently been developed to measure the fecal concentration of the inflammatory protein lactoferrin. Lactoferrin is elevated in IBD, but not in IBS or dysbiosis, and serves as an important diagnostic tool for chronic GI symptoms. Fecal lactoferrin assessment is a noninvasive screening for IBD (Crohn’s disease and ulcerative colitis).

Fecal Lysozyme

Lysozyme is secreted at sites of inflammation in the colonic mucosa. Lysozyme is a general marker for inflammation that is commonly elevated in the presence of dysbiosis. High fecal lysozyme levels may also accompany elevated fecal lactoferrin in IBD.

Fecal Mycology Culture

Identification of abnormal levels of specific yeast species in the stool is an important diagnostic step in therapeutic planning for the patient with chronic gastrointestinal and extra-gastrointestinal symptoms. Yeast sensitivities to a variety of prescriptive and natural agents are provided when yeast is cultured at any level. This provides the clinician with useful clinical information to help plan an appropriate treatment protocol. 

Fecal Ova & Parasites

Chronic gastrointestinal complaints are among the most common reasons that patients seek medical care. Fecal ova and parasite identification can identify a common underlying cause of chronic or acute GI symptoms.

Fecal Secretory IgA

The humoral immune status of the GI tract can be assessed by determining the fecal concentration of sIgA. The sIgA secreted by mucosal-associated lymphoid tissue (MALT) represents a pivotal and specific line of defense in the GI mucosa. As the principal immunoglobulin isotype present in mucosal secretions, sIgA plays an important role in controlling the intestinal milieu.

Fecal Short-Chain Fatty Acids (SCFAs)

SCFAs are the end product of the bacterial fermentation process of dietary soluble fiber by beneficial bacteria in the GI. SCFAs decrease inflammation, stimulate healing, and contribute to normal cell metabolism and differentiation.

Fecal Toxic Metals

Fecal toxic metal analysis provides important information about the potential for toxic metal burden. For many toxic metals, fecal (biliary) excretion is the primary natural route of elimination from the body. Fecal elemental analysis also provides a direct indication of dietary/oral exposure to toxic metals, and can be used to gauge mercury exposure from dental amalgams.

 

 

Hair Toxic Metals

Toxic metals may be 200-300 times more highly concentrated in hair than in blood or urine. Hair is the tissue of choice for detection of recent or ongoing exposure to elements such as arsenic, aluminum, cadmium, lead, antimony, and mercury.

Salivary Cortisol, DHEA

When cortisol is measured in four serial specimens (spanning from morning to bedtime on a day of typical stressor exposure), abnormalities of cortisol circadian release in response to pituitary ACTH may be assessed. The average of the two mid-day salivary dehydroepiandrosterone (DHEA) levels allows inspection for depressed formation from cholesterol in protracted adrenal dysfunction. Measurement of total sIgA allows detection of suppressed levels due to chronic elevated cortisol impact on the gut-associated immune tissue. Further evaluation of this impact is provided by inspecting for elevated anti-gliadin IgA.

Salivary Estrogen, Progesterone, Testosterone

Salivary estrogen, progesterone, and testosterone are convenient, noninvasive assays, measuring the free, bio-available form of these hormones. Salivary samples are collected at home, providing the opportunity to evaluate ovarian and adrenal function throughout their respective cycles, or to help account for mild fluctuations that occur in early menopause.  

Salivary Melatonin

Salivary melatonin is a convenient, noninvasive assay that is commonly used to assess the circadian secretion patterns over a complete light-dark cycle. This testing can reveal abnormal levels of melatonin that relate to various physical and psychological symptoms as well as premature acceleration of the body’s aging process.

Urinary Lactulose/Mannitol 

Oral challenge with a lactulose/mannitol sugar solution and measurement of urinary excretion identifies increased intestinal permeability and malabsorption. 

Urinary Amino Acids

Amino acid analyses aid in the diagnosis of: dietary protein adequacy and balance, gastrointestinal dysfunctions, forms of protein intolerance, nutritional deficiencies (vitamins, minerals), renal and hepatic dysfunction, psychiatric abnormalities, susceptibility to inflammatory response and oxidative stress, reduced detoxification capacity, susceptibility to occlusive arterial disease, and many inherent disorders of amino acid metabolism. Amino acid hepatic flux stimulates carcinogenic toxicant conjugation reactions. Amino acid analysis also shows phase II biotransformation substrates (glycine and sulfur amino acids) and status of glutathione precursor amino acids methionine and glycine. Neurotransmitter and neurotransmitter precursor assessment is provided by measurement of phenylalanine, tyrosine, tryptophan, glycine, taurine and aspartic and glutamic acids. Urinary hydroxylysine and hydroxyproline become elevated due to increased rates of bone resorption. Both specimens also show 3-methylhistadine rise in states of muscle catabolism. 

Urinary Bone Resorption

Urinary deoxypyridinoline (uDPD) assessment shows excretion of type I collagen peptide that is elevated when bone resorption increases to supply ionic calcium for critical cardiac and brain functions. Treatment efficacy can be monitored by repeat testing.

Urinary Challenge Detox Profiles

Challenge with caffeine, acetaminophen, and salicylic acid, with timed collection of urine and saliva, allows assessment of capacities for hepatic biotransformation phase I and II pathways, including CYP450 1A2, sulfation, glucuronidation, and glycination to meet toxin load demands. Measured compounds show the rate of Phase I caffeine clearance and percentage conversion of challenge compounds to oxidized, sulfated, glucuronidated, and glycinated products.

Urinary Estrogen Metabolites

Urinary 2-hydroxyestrone, 2-hydroxyestradiol and 16-hydroxyestrone and the calculated 2/16 ratio are measured to assess the function of the primary (hepatic CYP1A1) and secondary CYP1B1 estrogen clearance pathways. A 2/16 ratio less than 2 is associated with increased risk of developing estrogen sensitive cancers. Extensively studied diet and food supplement regimens can be used to increase CYP1A1 activity to raise the ratio.

Urinary Iodide

Iodine/Iodide is an essential element that is pivotal to normal function of the thyroid gland and the health and integrity of breast tissue. Iodine/Iodide intake has decreased significantly over the past thirty years as clinical symptoms have become more apparent. Iodine/Iodide sufficiency can be assessed by analysis of urinary iodide excretion with or without a loading protocol.

Urinary Mercapturic and D-glucaric Acids

Urinary D-glucaric acid, a byproduct of phase I detoxification, is a valuable indicator of chemical exposure or liver damage. Urinary mercapturic acids are direct end-product metabolites of conjugated xenobiotics. Combined assessment of the urinary levels of the two analytes provides valuable information about exposure to xenobiotics and liver disease, and the capability of the liver to eliminate toxins.

Urinary Neurotransmitter Metabolites

Urinary organic acid profiles show low or high levels of principal catabolites of norepinephrine and epinephrine (homovanilate) and dopamine (vanilmandelate).  These catecholamine metabolites reveal total body turnover that is depressed when the precursor amino acids phenylalanine and tyrosine are insufficient. Elevated levels found in patients with chronic stress response are a sign of accelerated utilization of the precursors that is antecedent to development of insufficiency. Similarly, the serotonin catabolite 5-hydroxyindolacetic acid reveals serotonin turnover and risk of tryptophan depletion. Simultaneous reporting of quinolinic acid and kynurenic acids allows inspection for inflammatory cytokine stimulation of glutamatergic neuronal activity.

Urinary Organic Acids

The full profile of organic acids in urine provides information on functional nutrient deficiencies, mitochondrial efficiency, methylation pathway cofactor sufficiencies, neurotransmitter metabolites, oxidant stress, detoxification and dysbiosis markers. A primary biochemical communication from the immune system to the CNS is assessed by measuring urinary interferon gamma-stimulated quinolinic acid (NMDA agonist) and kynurenic acid (NMDA antagonist). 

Urinary Oxidant Stress Markers

Urinary 8-hydroxy-2’-deoxyguanosine reveals oxidative damage to DNA. Urinary organic acid profiles include p-hydroxyphenyllactate (elevated in conditions of increased cell proliferation such as cancer or chronic organ stress); alpha-hydroxybutyrate (elevation shows increased glutathione demand); pyroglutamate (elevation reveals glutathione wasting); and sulfate (low levels indicate total body glutathione depletion).

Urinary Toxic Metals (provoked and unprovoked)

Urinary metal analysis is an invaluable tool for the diagnosis or confirmation of toxic element burden and monitoring of detoxification therapy. A challenge consisting of pre and post provocation testing is recommended for diagnosing the presence of toxic element burdens.