Advanced laboratory services

Overview 

The detection of inborn errors of metabolism (IEM) depends upon a high index of suspicion and coordinated access to expert laboratory services. Laboratory evaluation for IEM should be undertaken in all patients with suggestive history, examination, and/or abnormalities of routine laboratory tests.

Testing should be performed at the time of presentation or when symptoms are most pronounced because laboratory values may be normal when the patient is well. If at all possible, testing should be performed before treatment (or at least samples should be obtained before treatment, appropriately processed, and stored so that subsequent evaluation can be performed if indicated).

Patients with life-threatening illness should undergo concurrent evaluation for other conditions in the differential diagnosis (eg, sepsis, cardiac disease). Management of hypoglycemia, hyperammonemia, and seizures must be initiated promptly to prevent long-term sequelae.

A step-wise approach to evaluation is suggested, beginning with basic tests, before obtaining specialized metabolic investigations. The confirmatory diagnosis of most IEMs requires specialized testing that may include detection of abnormal metabolites in the plasma, urine, and/or CSF; assay of enzyme activity in skin, red blood cells, white blood cells, skeletal muscle, or liver; and/or chromosome or DNA analysis. 

Initial Evaluation  

At the time of the initial evaluation, blood and urine samples (and CSF if it is obtained) should be obtained (to the extent possible) for both basic tests and selected specialized tests, even though the specialized tests may not be necessary. This is because medical interventions may affect certain laboratory results that are necessary to establish the diagnosis (eg, administration of glucose containing IV fluids will affect the ability to detect hypoglycemia).

The initial evaluation of IEM includes: Complete blood count with differential, Blood gas analysis, Blood glucose, Serum ammonia, Electrolytes, blood urea nitrogen (BUN), creatinine, uric acid, Liver function tests: aminotransferases, bilirubin, prothrombin time, Examination of the urine, including color, odor, dipstick, and presence of ketones.

In addition, lactate dehydrogenase, aldolase, creatine kinase, and urine myoglobin should be obtained in patients who have complaints of muscle weakness, tenderness, cramping, atrophy, or exercise intolerance.

If possible, at the time of the initial evaluation, samples also should be obtained for the following specialized tests, that may be necessary depending on the results of the initial evaluation: Quantitative plasma amino acids, Acylcarnitine profile, Lactate, Qualitative urine organic acids .

Complete blood count (CBC) — Hematologic manifestations of IEM may involve any or all of the cell lines. The CBC also may provide a clue to sepsis, which may be the trigger for a metabolic crisis presenting manifestation of an IEM associated with increased risk of infection. 

Blood gas analysis — A blood gas analysis is used to detect acid-base disturbances. Metabolic acidosis with an increased anion gap is commonly associated with organic acidemias. Respiratory alkalosis is commonly seen in urea cycle disorders as a result of hyperammonemia.

Ammonia — The blood sample to measure ammonia concentration should be obtained from an artery or vein without using a tourniquet, placed on ice for transport to the laboratory, and analyzed immediately. If the plasma ammonia concentration is greater than 100 micromol/L (1.7 microgram/mL), the measurement should be repeated immediately. Significant elevations in ammonia (≥300 micromol/L [5.1 microgram/mL]) are most commonly associated with urea cycle disorders and certain organic acidemias (particularly propionic and methylmalonic acidemias). An elevated ammonia concentration (≥120 micromol/L [2.0 microgram/mL] in the newborn, and ≥80 micromol/L [1.4 microgram/mL] in older infants and children) is neurotoxic and must be treated immediately. The duration of hyperammonemia, rather than the peak, is predictive of poor developmental outcome in newborns.  

Electrolytes — Measurement of serum electrolytes is necessary to calculate the anion gap. A metabolic acidosis with an increased anion gap is commonly seen in organic acidemias. In addition, the finding of hyponatremia and hyperkalemia may provide a clue to salt-wasting.

Uric acid — Uric acid may be low in patients with defects of purine metabolism or molybdenum cofactor deficiency and increased in patients with Lesch-Nyhan disease or glycogen storage diseases (GSD).

Examination of urine — Several components of the urinalysis are helpful in the evaluation of the child with potential IEM:

- The presence or absence of ketones in the urine is helpful in determining the etiology of hypoglycemia.

- The urine pH is helpful in determining the cause of metabolic acidosis, if metabolic acidosis is present.

- Decreased urine specific gravity in a patient who is vomiting is suggestive of impaired ability to concentrate the urine, which is suggestive of renal tubular dysfunction (particularly when it occurs in conjunction with glucosuria and proteinuria). Renal tubular dysfunction occurs in a number of IEM.

- The presence of leukocyte esterase or nitrites on dipstick analysis is suggestive of urinary tract infection, which may be the precipitant for metabolic crisis, or the presenting manifestation of an IEM that has an associated increased risk of sepsis (eg, galactosemia, GSD type Ib [glucose-6-phosphatase deficiency, von Gierke disease]).

- The presence of reducing substances in the urine is a clue to certain IEM if the urine dipstick is negative for glucose. Children who have nonglucose reducing substances in the urine may have a carbohydrate intolerance disorder (eg, galactosemia, hereditary fructose intolerance) or an amino acid disorder. However, the absence of reducing substances in the urine does not exclude these disorders. False-positive tests for urine reducing substances in children may occur in children who have taken penicillins, salicylates, ascorbic acid, or drugs excreted as glucuronides.

Specialized Tests  

Specialized tests for IEM include quantitative plasma amino acids, qualitative urine organic acids, serum lactate, and acylcarnitine profile. Although these tests should only be performed as indicated by the clinical presentation and initial laboratory evaluation, the samples should be obtained at the time of acute presentation, if possible.

Plasma amino acids — Quantitative amino acid analysis in plasma or serum is used to confirm the diagnosis of urea cycle disorders and other disorders of amino acid metabolism. Quantitative plasma amino acid analysis is typically performed by high pressure liquid chromatography (HPLC), although tandem mass spectroscopy (MS/MS) can be used to measure amino acids. Most amino acids (except argininosuccinic acid and alloisoleucine) are present in the plasma within a normal range. Mild elevations of 5 to 10 percent above normal usually are not significant.

Amino acid analysis must be performed quantitatively rather than qualitatively. When a qualitative amino acid screen is performed by two-dimensional paper chromatography, elevations are reported only in groups of amino acids and specific disorders cannot be identified reliably.

Urine organic acids — Analysis of organic acids in urine is performed by gas chromatography/mass spectroscopy (GC/MS). A qualitative assay of these compounds is adequate because pathogenic organic acids (eg, methylmalonic or propionic acid) are not present in significant amounts in the urine of normal individuals.

Lactate and pyruvate — Lactate and pyruvate should be measured in arterial blood (tourniquet pressure and/or hemolysis may increase the lactate level erroneously), and transported on ice. For accurate measurement of pyruvate, the sample must be collected in perchlorate (or a similar media) to inactivate enzymes that degrade pyruvate.

Lactic acidosis caused by abnormal oxidative metabolism is a frequent finding in mitochondrial disorders (eg, disorders of oxidative phosphorylation), glycogen storage diseases, disorders of gluconeogenesis, and disorders of pyruvate metabolism. The ratio of lactate to pyruvate (normal value 10:1 to 20:1) may be helpful in differentiating among these conditions. The lactate-to-pyruvate ratio typically is high in mitochondrial disorders and in pyruvate carboxylase deficiency and normal or low in GSD and pyruvate dehydrogenase deficiency. However, exceptions occur, and the lactate-to-pyruvate ratio sometimes is normal in mitochondrial disorders. Elevated lactic acid also may be present in disorders of amino acid metabolism, organic acidemias, and fatty acid oxidation disorders (table).

Acylcarnitine profile — Analysis of acylcarnitine conjugates is performed by MS/MS and can be measured in a plasma sample or a filter-paper bloodspot. This test is used for the diagnosis of fatty acid oxidation disorders; it also may detect organic acidemias in which the acylcarnitine profile is abnormal (eg, propionic acidemia, isovaleric acidemia).