Automated HPLC screening of newborns for sickle cell anemia and other hemoglobinopathies.
Automated HPLC is used to test dried blood-spot specimens from newborns for hemoglobins (Hb) F, A, S, C, E, and D. We present the method and report on its performance determined during >4 years of testing 2.5 x 10(6) newborns. The method features automated derivation of presumptive phenotypes; quantitative quality control and proficiency testing; throughput of one specimen per minute; small sample volume; hemoglobin concentrations quantified with an interlaboratory CV of 14-18%; retention times with interlaboratory CV of <2% and matching, within +/- 0.03 min, of laboratories and reagent lots; control of peak resolution; 0.5% detection limit for Hb S and C, and 1.0% for Hb F, A, E, and D; few interferences; and negligible background and carryover. Shortcomings of the method are the absence of microplate barcode identification and the need for manually pipetting the sample eluate into the microplate.
Eastman
J W
JW
California Department of Health Services, Genetic Disease Laboratory, Berkeley, CA 94710, USA.
Wong
R
R
Liao
C L
CL
Morales
D R
DR
D000755:Anemia, Sickle Cell / Q000097:blood / Q000175:diagnosis*; D001322:Autoanalysis; D002851:Chromatography, High Pressure Liquid / Q000379:methods* / Q000706:statistics & numerical data; D005319:Fetal Hemoglobin / Q000032:analysis; D006441:Hemoglobin A / Q000032:analysis; D006444:Hemoglobin C / Q000032:analysis; D006446:Hemoglobin E / Q000032:analysis; D006451:Hemoglobin, Sickle / Q000032:analysis; D006453:Hemoglobinopathies / Q000097:blood / Q000175:diagnosis*; D006454:Hemoglobins / Q000032:analysis*; D006455:Hemoglobins, Abnormal / Q000032:analysis; D006801:Humans; D007231:Infant, Newborn; D015997:Neonatal Screening / Q000379:methods*; D012680:Sensitivity and Specificity
D006451:Hemoglobin, Sickle; D006454:Hemoglobins; D006455:Hemoglobins, Abnormal; C032001:hemoglobin D; D006444:Hemoglobin C; D006441:Hemoglobin A; D006446:Hemoglobin E; D005319:Fetal Hemoglobin