HbS Beta Research Articles

Submandibular TCD approach detects post-bulb ICA stenosis in children with sickle cell anemia

Transcranial Doppler (TCD) ultrasound is a procedure commonly used to screen individuals with the major hemoglobin S diseases, Hb SS and Hb S-beta(0), for significant stenoses in the circle of Willis. Flow velocities above 200 cm/s have been shown to identify patients at elevated risk for cerebral infarction. Among TCD's limitations is the inability to insonate the distal extracranial, petrous, and cavernous internal carotid artery (ICA) through the standard transtemporal approach. We extended the submandibular approach to include infra-siphon portions of the ICA. Using the extended submandibular approach to evaluate these portions of the ICA, we identified stenotic lesions in 4 patients with Hb SS disease out of a population of 131 children with Hb SS. Three of the 4 patients had no history of overt stroke or stroke-like symptoms. Neuroimaging confirmed the stenotic lesions, and also revealed watershed infarction as well as discrete areas of silent infarction. All 4 children had neuropsychological impairment. The submandibular approach, when added to a standard transcranial Doppler examination, may increase the sensitivity of this technique to identify important potential sources of cerebral infarction.

Mortality in Sickle Cell Patient Transitioning from Pediatric to Adult Program: 10 Years Grady Comprehensive Sickle Cell Center Experience.

Over the last three decades there has been improvement in survival in children with sickle cell disease. Overall survival from birth to age 18 of 86% and 95% has been reported in children with sickle cell anemia (HbSS) and sickle hemoglobin C (Hb SC) disease respectively. These encouraging results are secondary to initiation of preventive measures like newborn screening, penicillin prophylaxis, immunization and stroke prevention and other supportive therapies. Ballas (Blood 2004; 104 supl:Abstr 3743) reported high death rate for young adults with the disease. To determine the death rates and cause of death in our population, we did a retrospective review of our patients who had recently transitioned into adult care. Mortality rate and circumstances of death in a 10 year period of transition from pediatric to adult program in Georgia Comprehensive Sickle Cell Center at Grady Memorial Hospital between 1996 and 2006 were determined. Total number of deaths was obtained from the clinical database. Death, autopsy and medical records were reviewed to determine the cause and circumstances of death. Between 1996 and 2006, 387 young adults with sickle cell disease (HbSS, HbSC, HbS beta thalassemia) transitioned to the adult program at age 18. Twenty two (5.8%) patients died during their first 10 years of transition. Mean age at death was 23.4 (range 21.6–26) and male to female ratio was 1.2 to 1. Ten patients (45%) died due to chronic organ failure all due to chronic iron overload (end stage liver disease in 8 and congestive heart failure in 2). Eight patients (36%) died due to complications of acute vaso-occlusive crisis (sudden death in 4, acute chest/multi-organ failure in 4) while 4 patients (18%) died of non-sickle related causes. Three of four who died of sudden death has documented pulmonary hypertension. Our mortality rate is lower than those reported by Ballas with the most common cause of death being complications of iron overload in chronically transfused patients. Specific interventions targeted at improving management of iron overload need to be developed for the increasing numbers of patients on chronic transfusion transitioning into adult care.

Genetic polymorphisms associated with priapism in sickle cell disease

Priapism occurs in 30-45% of male patients with sickle cell disease (SCD), but the possible influence of genetic risk factors on the incidence of priapism is not well understood. We examined genetic polymorphisms in 199 unrelated, adult (>18 years), male patients with Hb SS and Hb Sbeta(0)-thalassaemia, 83 (42%) of whom reported a history of priapism. Candidate genes for association with priapism were identified based on their involvement in adhesion, coagulation, inflammation and cell signalling. Additionally, we examined genes involved in nitric oxide biology (NOS2, NOS3, SLC4A1), as well as polymorphisms in the klotho (KL) gene, which has previously been associated with priapism. Strong evidence of association was found for single nucleotide polymorphisms in transforming growth factor-beta receptor, type III (TGFBR3) (rs7526590; P = 0.00058), aquaporin (AQP1) (rs10244884; P = 0.00068), integrin alphav (ITGAV) (rs3768780; P = 0.00090), and the A1 subunit of coagulation factor XIII (F13A1) (hcv1860621; P = 0.00156). Associations with TGFBR3, AQP1, and ITGAV remained significant after adjusting for multiple testing, using the Benjamini-Hochberg procedure. Our data suggest that genes involved in the TGFbeta pathway, coagulation, cell adhesion and cell hydration pathways may be important in risk for priapism.

Proteomic Detection of Beta Thalassaemia Trait Using Electrospray Mass Spectrometry-Mass Spectrometry.

Introduction Mass spectrometry has proved a valuable tool in the unequivocal characterisation of haemoglobin variants detected during routine haemoglobinopathy screening using High Performance Liquid Chromatography (HPLC) and/or isoelectric focusing. We have recently described a rapid and specific electrospray mass spectrometry-mass spectrometry (MSMS) method, using proteomic mutation targeting, for simultaneous detection of the clinically significant haemoglobinopathies; haemoglobin (Hb)S, HbC, HbE, HbDPunjab and HbOArab. Here we report additional transitions within the 1min cycle time, using delta chains as a surrogate HbA2, with the objective of detecting beta thalassaemia trait.Method Blood samples are digested using trypsin, which results in specific cleavage of globin chains at peptide bonds adjacent to lysine and arginine residues, to produce a series of well-characterised peptides. There is strong amino acid sequence homology between the beta and delta chains of haemoglobin, however tryptic digestion produces series of 15 (T1–T15) and 16 (T1–T16) peptides, for beta and delta chains respectively, with differences in T2, T3, T5, T10, T12, T13 and T14. In this report we have investigated the diagnostic utility of T2: beta sequence SAVTALWGK, average mass 932.1 daltons, delta sequence TAVNALWGK, average mass 959.1 daltons. In multiple reaction monitoring mode the respective T2 [M+2H]2+ ions, m/z 466.8 and 480.3, are selected, fragmented, and the singly charged ions of the informative peptide fragments, VTALWGK and VNALWGK, detected.Results The percentage area ratio of delta/(beta+delta) was calculated and compared with the classical percentage HbA2 value obtained by HPLC. 66 samples with HbA2 values within our normal range (mean 2.7%, range 1.8–3.4), and 58 with HbA2 values indicative of beta thalassaemia trait (5.2%, 4.2–7.9) were analysed by MSMS and the corresponding delta/(beta+delta) ratios were 1.7%, 0.9–2.3 and 3.4%, 2.5–6.0. In addition, in 2 further HbS beta thalassaemia zero compound heterozygote samples confirmed by beta gene sequencing the ratios were 4.1% and 3.7%.Conclusion This relatively small study provides evidence that beta thalassaemia trait may be diagnosed by MSMS and represents another step towards comprehensive MSMS population screening for clinically significant haemoglobinopathies.

Microalbuminuria as a predictor of early glomerular injury in children with sickle cell disease.

A cross sectional study was carried out to determine the prevalence of microalbuminuria in the pediatric patients with sickle cell disease. The study was carried out on 64 pediatric patients aged less than 14 years with documented HbSS, HbAS and HbS beta thalassemia, Microalbuminuria was estimated using single radial immuno diffusion technique. Majority of the study subjects were of HbSS type. 38.5% had symptoms for > 2 years. 18.8% of the study population had significant microalbuminuria (19.2% of SS type and 18.8% of Hb AS types). Microalbuminuria excretion was significantly more in patients > 9 years of age as compared to young patients (p < 0.05). Mean serum creatinine levels did not show any significant difference in the various study groups. Microalbuminuria estimation is a very important clinical marker of preclinical glomerular damage in patients with sickle cell disease. It estimation would help in the early detection of such patients and prompt initiation of therapy.

Characterization of pica prevalence among patients with sickle cell disease.

To determine the prevalence of pica and its characteristics among children with sickle cell disease. Retrospective, observational study. An urban, ambulatory care, interdisciplinary center. The medical records of all 480 patients who visited the center from March 1, 1998, to June 30, 1999, were reviewed. Patients were excluded for history of stroke, long-term transfusions, pregnancy, acute illness, or age younger than 3 years. Sex, age, weight, height, Tanner stage, complete blood cell count, sickle cell genotype, pica history, and levels of iron, zinc, lead, and fetal hemoglobin (Hb). Of 395 study patients, 134 (33.9%) reported pica. Ingested items included paper, foam, and powders. There was a significantly higher prevalence of pica among patients homozygous for Hb S (Hb SS, sickle cell anemia) compared with the combined group of double heterozygous patients with Hb SC, Hb SD, and Hb Sbeta thallasemia (Sbeta(+)or Sbeta(0)) (35.6% vs 25.5%; P =.03). Within genotype, mean Hb levels were significantly lower and reticulocyte counts were significantly higher in the patients with pica. Overall, the mean age of patients with pica was significantly lower; however, the prevalence was 23.3% (27/116) among those aged 10.0 to 14.9 years and 14.8% (8/54) among those aged 15.0 to 19.0 years. Within age groups, patients with pica weighed significantly less. Pica appeared to have an unusually high prevalence in patients with sickle cell disease and a correlation with lower Hb levels. It is unclear whether pica is a specific marker of disease severity, because our review did not show a relationship to increased number and duration of hospitalizations. The association between pica and low body weight suggests a nutritional effect on its prevalence.

Acute Chest Syndrome in Adults With Sickle Cell Disease

Acute chest syndrome (ACS) is a frequent and potentially severe pulmonary illness in sickle cell disease (SCD). The aim of the study was to report the clinical features and outcome of consecutive ACS episodes in adult patients in a French SCD center. All patients were treated according to an uniform therapeutic protocol applying transfusion only in the more severe clinical form of ACS. There were 107 consecutive episodes in 77 adult patients (mean age, 29 +/- 7 years; 78% hemoglobin [Hb] SS; 14% Hb SC; and 8% Hb Sbeta + thalassemia) over a 6-year period. Seventy-eight percent of our patients had an associated vaso-occlusive crisis that preceded the chest signs in half of the cases. Comparison between acute and baseline levels showed a statistically significant difference in Hb levels (drop of 1.6 to 2. 25 g/dL depending on Hb genotype), WBC count (increase of 9.2 +/- 8. 3 x 10(9)/L); platelet count (increase of 67 +/- 209 x 10(9)/L); and lactate dehydrogenase values (increase of 358 +/- 775 IU/L) in ACS patients. Hypercapnia was detected in 42% of patients without sign of narcotic abuse. We identified a high percentage of alveolar macrophages containing fat droplets in 31 of 43 (77%) patients who underwent BAL. Bacterial culture findings were almost always negative, but were performed after starting antibiotic therapy that was administered in 96 episodes. Transfusion was required in 50 of 107 ACS events (47%). Five patients died, and all were transfused. These results confirm that fat embolism is probably a frequent mechanism of ACS in adult patients. However, fat embolism was not associated with a more severe clinical course, suggesting that bronchoscopy and BAL have little impact on the management of these patients. Restricting transfusion to the most severe ACS cases does not seem to increase the mortality rate.

Free energy simulations of axial contacts in sickle-cell hemoglobin

Molecular dynamics simulations have been used to investigate the thermodynamic stability of axial contacts in sickle-cell hemoglobin (HbS). Free energy changes were evaluated for the point mutation beta 121 Glu --> Gln in the axial contact region of HbS crystals. The calculations predict a free energy change of-3.6 kcal/mol per contact for the mutation, which is in qualitative agreement with experimental observations of aggravated sickling found in the double mutant Hb D Los Angeles (beta 6 Glu --> Val. beta 121 Glu --> Gln) relative to HbS (beta 6 Glu --> Val). The beta 121 Glu is sequestered in a salt link with beta 17 Lys located on the same polypeptide chain, making the Glu interactions with its surroundings similar in aggregates and individual hemoglobins. Due to this cancellation of the large electrostatic Glu contributions, the weak nonspecific interactions between the Gln and the neighboring polypeptide chain are the main contributing factor to the enhanced aggregation of Hb D Los Angeles relative to HbS. Together with the previous study of the lateral contact [K. Kuczera et al. (1990) Proceedings of the National Academy of Science USA, Vol. 87, pp, 8481-8485], the present results provide a more complete picture of the forces driving the sickling aggregation. A comparison of different treatments of internal flexibility in free energy simulations and analysis of rate of convergence of the different calculated properties has also been performed.

Molecular characterization of β-thalassemia mutations in Guadeloupe

In order to perform genetic counselling and prenatal diagnosis of Hb-S-beta-thalassemia disease and beta-thalassemia, we have delineated the spectrum of beta-thalassemia alleles in the Guadeloupean population. A sample of 63 unrelated families was analyzed including 70 beta-thalassemia carriers, 52 Hb-S-beta-thalassemia, and 8 patients with different beta-thalassemic hemoglobinopathies. Among the eleven mutations identified, four of them [-29 (A --> G), IVS-I-5 (G --> A), IVS-II-1 (G --> A), and IVS-I-5 (G --> C)] account for 77.6% of the beta-thalassemia chromosomes present in the studied families. The seven other variants, CD 24 (T --> A), IVS-I-2 (T --> C), Poly A (T --> C), -88 (C --> T), IVS- 11-849 (A --> G), Hb E, and Hb Lepore are less frequent. As a result, Hb S-beta+-thalassemia type 1 (low Hb A values: 5-15%) together with Hb S-beta(omicron)-thalassemia phenotypes are as frequent as Hb S-beta+-thalassemia type 2 (high Hb A values: 20-30%) in the Guadeloupean population. Patients with Hb S-beta+-thalassemia type 2 have milder hematological manifestations of the disease compared to patients with Hb S-beta(omicron)-thalassemia and Hb S-beta+-thalassemia type 1. This first report on the type and nature of beta-thalassemia mutations in Guadeloupe shows that prenatal diagnosis of Hb S-beta-thalassemia and beta-thalassemia should be feasible by direct detection of point mutation in most cases.

Molecular characterization of beta-thalassemia mutations in Guadeloupe.

In order to perform genetic counselling and prenatal diagnosis of Hb-S-beta-thalassemia disease and beta-thalassemia, we have delineated the spectrum of beta-thalassemia alleles in the Guadeloupean population. A sample of 63 unrelated families was analyzed including 70 beta-thalassemia carriers, 52 Hb-S-beta-thalassemia, and 8 patients with different beta-thalassemic hemoglobinopathies. Among the eleven mutations identified, four of them [-29 (A --> G), IVS-I-5 (G --> A), IVS-II-1 (G --> A), and IVS-I-5 (G --> C)] account for 77.6% of the beta-thalassemia chromosomes present in the studied families. The seven other variants, CD 24 (T --> A), IVS-I-2 (T --> C), Poly A (T --> C), -88 (C --> T), IVS- 11-849 (A --> G), Hb E, and Hb Lepore are less frequent. As a result, Hb S-beta+-thalassemia type 1 (low Hb A values: 5-15%) together with Hb S-beta(omicron)-thalassemia phenotypes are as frequent as Hb S-beta+-thalassemia type 2 (high Hb A values: 20-30%) in the Guadeloupean population. Patients with Hb S-beta+-thalassemia type 2 have milder hematological manifestations of the disease compared to patients with Hb S-beta(omicron)-thalassemia and Hb S-beta+-thalassemia type 1. This first report on the type and nature of beta-thalassemia mutations in Guadeloupe shows that prenatal diagnosis of Hb S-beta-thalassemia and beta-thalassemia should be feasible by direct detection of point mutation in most cases.

Genetic heterogeneity of beta-thalassemia in southeast Sicily.

In this study we have defined the spectrum of the beta-thalassemia mutations, the beta-thalassemia haplotypes, and the genotype-to-phenotype correlations in a large number of patients with different beta-thalassemia conditions. Seventeen different beta-thalassemia mutations were detected which included one chromosome each with Hb Dhonburi and Hb Lepore. Five alleles, namely, codon 39 (C-->T), IVS-I-110 (G-->A), IVS-I-6 (T-->C), IVS-II-745 (C-->G), and IVS-I-1 (G-->A), account for 90% of all beta-thalassemia mutations in 846 thalassemic chromosomes studied. Haplotyping for a large number of subjects showed that the five common mutations are linked to a few haplotypes. The presence of milder mutations, mainly IVS-I-6 (T C), in about 19% of our patients explains some of the clinical variables. Among the 37 patients with thalassemia of intermediate severity, only 6 were homozygous or compound heterozygous for two severe alleles. The type of beta-thalassemia is the main factor responsible for differences in the phenotypic expression of the disease in patients with Hb S-beta-thalassemia; patients with Hb S-beta(+)-thalassemia are less severely affected than those with Hb S-beta(0)-thalassemia. The five most frequent mutations have comparable distributions all over Sicily.

Sickle cell rheology is determined by polymer fraction--not cell morphology.

Sickle cell disease pathophysiology is mediated by acute and chronic impairment of cell flexibility due to the formation of intracellular sickle hemoglobin (Hb S) polymer as cells are partially deoxygenated in the microcirculation. We have recently developed a method to measure the relationship between the formation of intracellular polymerized Hb S and cell filtration. In this study, we have used this method to examine whether sickle cell morphology, independent of Hb S polymer fraction, had an effect on cell rheology. We primarily use sickle trait (AS) and Hb S-beta(+)-thalassemia (S-beta(+)-thal) erythrocytes with low hemoglobin F levels, which have normal membranes and few or no dense cells, to remove these confounding effects. We find that the relationship between filtration and the percentages of each "type" of morphological deformation of AS erythrocytes was different from that of the S-beta(+)-thal erythrocytes. In addition, we find that while the filtration of AS erythrocytes as a function of oxygen saturation was similar, whether measured during deoxygenation or reoxygenation, the relationship between the percentages of each type of deformed erythrocyte and oxygen saturation demonstrated hysteresis during oxygenation-deoxygenation experiments. Transmission electron microscopy, for both elongated and irregularly shaped cells, showed that similarly distorted cells could have very different amounts and alignment of polymer. These results suggests that cell morphology per se is not strongly related to filtration, whereas calculated intracellular Hb S polymer fraction predicts loss of filtration of AS and S-beta(+)-thal erythrocytes well. Measured or calculated polymer fraction values would appear to be a better parameter for the study of sickle cell disease pathophysiology and response to treatment than cell morphology studies.

The relative levels of βA and βS mRNAs in Hb S heterozygotes and in patients with Hb S‐β+ ‐thalassaemia or Hb S‐β+ ‐HPFH combinations

We have used a quantitative reverse transcription/polymerase chain reaction (RT/PCR) procedure to evaluate the relative amounts of beta A and beta S mRNA transcripts in eight subjects with a simple Hb S heterozygosity, in six with Hb S-beta(+)-thalassaemia (thal), and in three individuals with Hb S-beta(+)-HPFH [hereditary persistence of fetal haemoglobin (Hb)] [two with the Atlanta type and one with the G gamma-202 (C-->G) substitution]. A balanced synthesis of beta A and beta S mRNAs was observed in all Hb S heterozygotes, whereas the beta A mRNA levels were reduced to approximately 16% of that of the beta S mRNA in the six Hb S-beta(+)-thal compound heterozygotes, to approximately 43% in the two subjects with Hb S-beta(+)-HPFH (Atlanta type), and to 23.8% in the one individual with Hb S-beta(+)-HPFH [G gamma-202 (C-->G) substitution]. The higher Hb A versus Hb S levels observed in all groups of the patients studied, further confirm a post-translational control mechanism in determining the levels of Hb A and Hb S in the peripheral blood of these individuals. The procedure described here provides an accurate and easy method for studying the relative expression of particular globin genes at the transcriptional level in patients with various haemoglobinopathies.

A mild type of Hb S‐β+‐thalassemia [‐92(C→T)] in a sicilian family

Hematological data are presented for an adult Sicilian patient with a mild Hb S-beta(+)-thalassemia caused by a C-->T mutation at position -92 of the beta promoter. This mutation was identified by sequencing of amplified DNA and was confirmed by dot-blot analysis with specific probes. A comparison of levels of Hb S and Hb A in Hb S-beta-thalassemia patients with different beta-thalassemia alleles showed great variations; the highest level of Hb A (45%) was recorded in the patient with Hb S-beta(+)-thalassemia [-92(C-->T)] and the lowest (approximately 13%) in patients with Hb S-beta(+)-thalassemia [IVS-II-745 (C-->G)]. Clinical severity is directly related to the level of Hb A present.

Molecular characterization of Hb S(C) beta-thalassemia in American blacks.

An extension of previous reports describing the molecular defects and hematological abnormalities in black patients with Hb S(C) beta-thalassemia living in the Southeastern United States is presented. As many as 58 patients with Hb S-beta(+)-thalassemia, 16 with Hb C-beta(+)-thalassemia and 12 with Hb S-beta(0) -thalassemia have been studied. Patients with Hb S(C) beta(+)-thalassemia type 2 (high Hb A values) were most common; the thalassemia was due to mutations in the promoter of the beta-globin gene [-88 (C----T) and -29 (A----G)] or at the polyadenylation signal (T----C). Two patients with lower Hb A values (type 1) carried a mutation in the first intron of the beta-globin gene (IVS-1-5: G----T). The simultaneous presence of an alpha-thalassemia -2(-alpha/) resulted in some modifications of the hematological parameters, but had a minimal effect on the clinical condition. Patients with Hb S-beta (0) thalassemia had lower hemoglobin values, lower number of red blood cells, and lower MCHC values and suffered more frequently from complications than the patients with Hb S-beta(+)-thalassemia. A total of 17 different beta-thalassemia mutations were observed in 128 chromosomes; two mild beta(+)-thalassemia mutations [-88(C----T) and -29(A----G)] account for more than 80% of the thalassemic chromosomes.

Hemoglobinopathies Among the Gond Tribal Groups of Central India; Interaction of α- and β-Thalassemia with β Chain Variants

We have investigated the frequencies and types of alpha-thal, beta-thal, and Hb variants among nearly 200 inhabitants of villages in the Mandla and Jabalpur districts of Madhya Pradesh in Central India. Over 85% were tribals of the Gond group. alpha-Thal, as -alpha 3.7/and -alpha 4.2/, and the nondeletional Koya Dora mutation were present at the combined frequency of 0.54. There were indications for the presence of other nondeletional types of alpha-thal. alpha-Globin gene triplications were not observed. Four of the six beta-thal alleles observed were in the tribal groups; two (G----C at codon 30 and G----A at IVS-I-1) were found for the first time. The simultaneous presence of an alpha-thal (-alpha/alpha alpha or -alpha/-alpha) greatly improved the clinical and hematological condition of the patients with Hb S-beta(+)-thal (IVS-I-5; G----C). The lower frequency of alpha-thal among the beta-thal heterozygotes (f = 0.32) may indicate that some of the beta-thal alleles in the tribal populations originated from an outside source. Forty-one subjects had SS; all but one had beta S with haplotype #31, while one chromosome had haplotype #17. The presence of an alpha-thal-2 (f = 0.53) in the SS patients did not affect hematological data. The Hb F levels varied between 7.5% and 42.5% with high G gamma values. No difference in Hb F level between males and females was observed. Lower Hb F levels were present in 10 SS patients with an alpha-thal-2 homozygosity (average 16% versus 23.5% for eight SS patients with alpha alpha/alpha alpha) suggesting a decreased formation of alpha gamma dimers in severe alpha chain deficiency. Several younger SS patients (less than 10 years) also had high Hb F levels (32-42%). Variations in the sequence at -530 of the beta-globin gene; i.e. in the so-called silencer sequence, were present in all beta S chromosomes with haplotype #31, but were not considered important for understanding the variability in the Hb F level. gamma-Globin gene deletions (gamma-thal) and triplications were not observed.

Incidence of overt and covert neurological damage in children with sickle cell disease.

We studied 25 patients with sickle cell disease (15 with HbSS; 8 with HbSC and 2 with HbS beta degree) using neurological examination as a marker of overt and computed tomographic (CT) scan as a marker of covert neurological damage. Of the 25 patients, 5 (20%) had evidence of cerebral infarcts on CT scan, but only 4 of these 5 patients had neurological signs. Neurological problems were only seen in patients with HbSS. We could not identify any haematological risk factors, but in this small group of HbSS patients a crisis rate greater than 1 per year of follow up was associated with a significant risk of neurological damage.

Newborn Screening for Hemoglobin Abnormalities: A Comparison of Methods

Peripheral blood from 1,000 newborn infants of black (641) and Southeast Asian (359) ancestry were screened for hemoglobin variants. Results obtained from the combination of cellulose acetate (CAC) and citrate agar (CAG) electrophoresis were compared with isoelectric focusing (IEF) electrophoresis. There was complete agreement between the two methods on assignment of Hb S trait, Hb C trait, Hb E trait, and homozygous Hb E. IEF identified small amounts of Hb A in two newborn infants with Hb S-beta + thalassemia; the CAC and CAG electrophoretic patterns were indistinguishable from sickle cell anemia. One hundred twenty newborn infants with Hb Bart's were detected by IEF; 51 of these were found on CAC. Although IEF was more sensitive in detecting small amounts of hemoglobin, it is not clear if the improvement in detection warrants adopting this form of electrophoresis for routine screening of newborn infants.