Test ID: CDG
Carbohydrate Deficient Transferrin for Congenital Disorders of Glycosylation, Serum

Screening for congenital disorders of glycosylation

Results are reported as the mono-oligosaccharide/di-oligosaccharide transferrin ratio, the a-oligosaccharide/di-oligosaccharide transferrin ratio, the tri-sialo/di-oligosaccharide transferrin ratio, the apolipoprotein CIII-1/apolipoprotein CIII-2 ratio and the apolipoprotein CIII-0/apolipoprotein CIII-2 ratio.

• Informed Consent for Genetic Testing

Affinity Chromatography-Mass Spectrometry (MS)

Collection Container/Tube:

Preferred: Red top

Acceptable: Serum gel

Submission Container/Tube: Plastic vial

Specimen Volume: 0.1 mL

Additional Information: 

1. Patient's age is required.

2. Reason for referral is required.

3. This test is for congenital disorders of glycosylation. If the ordering physician is looking for evaluation of alcohol abuse, order CDTA/82425 Carbohydrate Deficient Transferrin, Adult, Serum.

Forms:

1. New York Clients-Informed consent is required. Please document on the request form or electronic order that a copy is on file. An Informed Consent for Genetic Testing (Supply T576) is available in Special Instructions.

 2. If not ordering electronically, submit a Biochemical Genetics Request Form (Supply T439) with the specimen.

Congenital disorders of glycosylation (CDG), formerly known as carbohydrate-deficient glycoprotein syndrome, are a group of more than 45 inherited metabolic disorders affecting several steps of the pathway involved in the glycosylation of proteins. CDG are classified into 2 groups. Type I CDG is characterized by defects in the assembly or transfer of the dolichol-linked glycan, while type II involves processing defects of the glycan. Apolipoprotein CIII (Apo-CIII) isoforms, a protein with a single core 1 mucin type O-glycosylate protein, is a complementary evaluation for the CDG type II profile. This analysis will evaluate mucin type O-glycosylation, a defect that happens in the Golgi apparatus, and will change the ratios, increasing the asialo or monoisalo forms and decreasing the fully sialilate (disialo) forms. In young children (>1 month) and in liver disease, the Apo-CIII2 may be increased. Children younger than 6 months, and clinically suspected of having ATP6V0A2-CDG, may have normal transferrin profile with abnormal Apo-CIII profile.

CDG typically present as multisystemic disorders with a broad clinical spectrum including: developmental delay, hypotonia, with or without neurological abnormalities, abnormal magnetic resonance imaging findings, skin manifestations, and coagulopathy. There is considerable variation in the severity of this group of diseases, ranging from hydrops fetalis to a mild presentation in adults. In some subtypes (Ib, in particular), intelligence is not compromised. CDG should be suspected in all patients with developmental delay, unexplained liver dysfunction, stroke-like episodes, unexplained hypoglycemia, and liver disease. Abnormal subcutaneous fat distribution and chronic diarrhea each may or may not be present. The differential diagnosis of abnormal transferrin patterns also includes liver disease not related to CDG including uncontrolled galactosemia, hereditary fructose intolerance in acute crisis, and liver disease of unexplained etiology.

Transferrin and apolipoprotein CIII isoform analysis test is the initial screening test for CDG. The results of the transferrin and apolipoprotein CIII isoform analysis should be correlated with the clinical presentation to determine the most appropriate testing strategy including enzyme, molecular, and research-based testing. If either CDG-Ia or CDG-Ib are suspected, enzymatic analysis for phosphomannomutase and phosphomannose isomerase in leukocytes (PMMIL/89656 Phosphomannomutase [PMM] and Phosphomannose Isomerase [PMI], Leukocytes) or fibroblasts (PMMIF/89657 Phosphomannomutase [PMM] and Phosphomannose Isomerase [PMI], Fibroblasts) should be performed.

Positive test results could be due to a genetic or nongenetic condition; additional confirmatory testing is required.

Results are reported as the mono-oligosaccharide/di-oligosaccharide transferrin ratio, the a-oligosaccharide/di-oligosaccharide transferrin ratio, the tri-sialo/di-oligosaccharide transferrin ratio, and the apolipoprotein CIII-1/apolipoprotein CIII-2 ratio, and the apolipoprotein CIII-0/apolipoprotein CIII-2 ratio. The report will include the quantitative results and an interpretation.

The congenital disorders of glycosylation (CDG) profiles are categorized in 4 types: 

-CDG type I profile. Mono-oligosaccharide/di-oligosaccharide transferrin ratio, and/or the a-oligosaccharide/di-oligosaccharide transferrin ratio are abnormal. This group should have the apoliprotein C-III profile within the normal ranges, because the Golgi system is not affected in CDG type I.

-CDG type II profile. The tri-sialo/di-oligosaccharide transferrin ratio is abnormal. In this category, the apolipoprotein C-III profile will have 2 scenarios:

   - The apolipoprotein CIII-1/apolipoprotein CIII-2 ratio and/or the apolipoprotein CIII-0/apolipoprotein CIII-2 ratio will be abnormal when the defect is most likely glycan processing in the Golgi apparatus, therefore the CDG defect is likely.

   - The apolipoprotein CIII-1/apolipoprotein CIII-2 ratio and/or the apolipoprotein CIII-0/apolipoprotein CIII-2 ratio are normal, in this case most likely the defects do not involve the Golgi system, thus the molecular defect is different in this case.

-CDG mixed type profile (type I and II together). In this type of profile one can have abnormal tri-sialo/di-oligosaccharide transferrin ratio with the mono-oligosaccharide/di-oligosaccharide transferrin ratio and/or the a-oligosaccharide/di-oligosaccharide transferrin ratio abnormal, and may have the apolipoprotein CIII-1/apolipoprotein CIII-2 ratio and the apolipoprotein CIII-0/apolipoprotein CIII-2 ratio normal or abnormal, depending if the defects involve Golgi apparatus.

When the profile cannot be categorized following the above classification, all the abnormal transferrin and/or Apo-CIII species will be reported descriptively according to the molecular mass stating the possible structures.

Reports of abnormal results will include recommendations for additional biochemical and molecular genetic studies to more precisely identify the correct form of CDG. Treatment options, the name and telephone number of contacts who may provide studies at Mayo Clinic or elsewhere, and a telephone number for one of the laboratory directors (if the referring physician has additional questions) will be provided.

Other conditions such as acute crisis of hereditary fructose intolerance, metabolically decompensate galactosemia, and acute liver disease may have a congenital disorders of glycosylation (CDG) profile that is indistinguishable from any other true CDG type I cases. Relevant clinical information and the indication for the analysis should be provided with the specimen, in particular for nonpediatric patients.

1. Jaeken J, Matthijs G: Congenital disorders of glycosylation: a rapidly expanding disease family. Annu Rev Genomics Hum Genet 2007;8:261-278

2. Freeze HH: Congenital disorders of glycosylation: CDG-I, CDG II, and beyond. Curr Mol Med 2007;7:389-396

3. Patterson MC: Screening for "prelysosomal disorders": carbohydrate-deficient glycoprotein syndromes. J Child Neurol 1999;14(Suppl 1):S16-S22

4. Stibler H, Jaeken J: Carbohydrate deficient serum transferrin in a new systemic hereditary syndrome. Arch Dis Child 1990;65:107-111

5. Freeze HH: Disorders in protein glycosylation and potential therapy - tip of an iceberg. J Pediatr 1998;133:593-600

6. Stibler H, Borg S: Evidence of a reduced sialic acid content in serum transferrin in male alcoholics. Alcohol Clin Exp Res 1981;5:545-549

Samples are prepared by diluting serum (25 microliters) in water (100 microliters). Ten microliters of the diluted sample is then injected into a 200 microliters/minute flow of pH 7.4 phosphate buffer saline (PBS) and retained on an immunoaffinity column that is composed of 60% antihuman apolipoprotein CIII antibody and 40% antihuman transferrin antibody bound to POROS  20 AL media. The immunoaffinity column is then washed for 2 minutes with PBS. Following the 2-minute wash, captured proteins are eluted to a C4 column at 200 microliters/minute with pH 2.5 100mM glycine/2% acetic acid buffer for 2.5 minutes. The C4 column is then washed for 1 minute with water/methanol/glacial acetic acid (97/2/1) at 200 microliters/minute to remove signal-suppressing salts. The proteins are then eluted from the C4 column with methanol/water/glacial acetic acid/TFA (94.5/5/0.5/0.04) at 200 microliters/minute and introduced into an API 4000 tandem mass spectrometer equipped with a Turbo V source configured for electrospray ionization. The mass spectrometer is operated in positive Q1 scan mode with 2 scan ranges; m/z 1090-2000 for apolipoprotein CIII and m/z 2000-3000 for transferrin. The total analysis time is 8 minutes, including column equilibration. Relative quantitation of carbohydrate deficient transferrin and apolipoprotein CIII is achieved by comparing glycoform ratios in each protein.(Lacey JM, Bergen R, Magera MJ, et al: Rapid determination of transferrin isoforms by immunoaffinity liquid chromatography and electrospray mass spectrometry. Clin Chem 2001;47:513-518)

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