Diagnosis of congenital disorders of glycosylation type-I using protein chip technology.
A method for the diagnosis of the congenital disorders of glycosylation type I (CDG-I) by SELDI-TOF-MS of serum transferrin immunocaptured on protein chip arrays is described. The underglycosylation of glycoproteins in CDG-I produces glycoforms of transferrin with masses lower than that of the normal fully glycosylated transferrin. Immobilisation of antitransferrin antibodies on reactive-surface protein chip arrays (RS100) selectively enriched transferrin by at least 100-fold and allowed the detection of patterns of transferrin glycoforms by SELDI-TOF-MS using approximately 0.3 microL of serum/plasma. Abnormal patterns of immunocaptured transferrin were detected in patients with known defects in glycosylation (CDG-Ia, CDG-Ib, CDG-Ic, CDG-If and CDG-Ih) and in patients in whom the basic defect has not yet been identified (CDG-Ix). The correction of the N-glycosylation defect in a patient with CDG-Ib after mannose therapy was readily detected. A patient who had an abnormal transferrin profile by IEF but a normal profile by SELDI-TOF-MS analysis was shown to have an amino acid polymorphism by sequencing transferrin by quadrupole-TOF MS. Complete agreement was obtained between analysis of immunocaptured transferrin by SELDI-TOF-MS and the IEF profile of transferrin, the clinical severity of the disease and the levels of aspartylglucosaminidase activity (a surrogate marker for the diagnosis of CDG-I). SELDI-TOF-MS of transferrin immunocaptured on protein chip arrays is a highly sensitive diagnostic method for CDG-I, which could be fully automated using microtitre plates and robotics.
Biochemistry, Endocrinology and Metabolism Unit, UCL Institute of Child Health, London, UK.
Kevin
K
Mills
Philippa
P
Mills
Marie
M
Jackson
Viki
V
Worthington
Clare
C
Beesley
Anúska
A
Mann
Peter
P
Clayton
Stephanie
S
Grunewald
Geoffrey
G
Keir
Liz
L
Young
James
J
Langridge
Nasi
N
Mian
Bryan
B
Winchester
D000595:Amino Acid Sequence; D002239:Carbohydrate Metabolism, Inborn Errors / Q000097:blood* / Q000175:diagnosis* / Q000235:genetics; D015180:Electrophoresis, Gel, Two-Dimensional; D006023:Glycoproteins / Q000097:blood; D006031:Glycosylation; D006801:Humans; D008969:Molecular Sequence Data; D011110:Polymorphism, Genetic; D040081:Protein Array Analysis / Q000379:methods; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014168:Transferrin / Q000032:analysis / Q000235:genetics / Q000378:metabolism
D006023:Glycoproteins; D014168:Transferrin