Polimorfismo genético da glicose-6-fosfato desidrogenase na população da região de Araraquara, Estado de São Paulo / Genetic polymorphism of glucose-6-phosphate dehydrogenase in the population of Araraquara, State of São Paulo

The most important role played by the enzyme Glucose-6-Phosphate Dehydrogenase (G6PD) in erythrocyte metabolism is in generating energy and reducing power used to protect the cell against oxidative attack. G6PD deficiency is the erythroenzymopathy that most frequently causes hemolytic anemia, and more than 130 molecular variants have already been identified. The aim of this study was to analyze the genetic mutations in the G6PD-deficient adult males in the population of the region of Araraquara, São Paulo State. Out of 5087 male blood donors, 89 were deficient for G6PD, as confirmed by assaying the enzyme activity and electrophoresis on cellulose acetate. Thus, a frequency of 1.75% of G6PD-deficient patients was found, this value being similar to other investigations in São Paulo state. Molecular analysis was performed by amplification of genomic DNA with specific primers and digestion with restriction enzymes. In 96.6% of the patients, the G6PD A¯ variant was observed, with mutations at residues 376(A-G) and 202(G-A). Mean G6PD specific activity among the patients was 1.31 IU.g Hb-1.min-1 at 37ºC, that is 10.8% of the normal activity of the G6PD B enzyme. The variant forms G6PD A¯680(G-T) and 968(T-C) were not found. In 3.4% of the deficient individuals, the G6PD Mediterranean variant was found, with a mutation at 563(C-T). In these cases,mean enzymatic activity was 0.25 IU.g Hb-1.min-1 at 37ºC, or 2.1% of the enzymatic activity of G6PD B. Theuse of traditional techniques, allied to the identification of the different molecular variants, is important for the understanding of the structural and functional properties and hemolytic behavior of the red blood cells of the patient...

A importância da enzima Glicose-6-fosfato desidrogenase (G6PD) no metabolismo eritrocitário está na obtenção de energia calórica e redutora para a proteção celular contra agressões oxidativas. A deficiência de G6PD é a eritroenzimopatia que causa mais frequentemente anemia hemolítica, com mais de 130 variantes moleculares identificadas. O objetivo deste estudo foi realizar a análise molecular da deficiência de G6PD em uma população masculina adulta da região de Araraquara, SP, para a identificação das mutações genéticas. Nos 5087 doadores de sangue do sexo masculino pesquisados, foram encontrados 89 deficientes de G6PD, confirmados pela determinação da atividade enzimática e eletroforese em acetato de celulose, com frequência de 1,75%, valores semelhantes aos encontrados por outros pesquisadores no Estado de São Paulo. A análise molecular realizada pela amplificação do DNA genômico com iniciadores específicos e digestão com enzimas de restrição, demonstrou que 96,6% dos deficientes apresentaram a variante G6PD A¯, com as mutações 376(A-G) e 202(G-A) e atividade enzimática média de 1,31 UI.g de Hb-1.min-1 a 37ºC, correspondendo a 10,8% da atividade enzimática da enzima normal G6PD B. Não foram encontradas as formas variantes G6PD A¯ 680(G-T) e 968(T-C). Em 3,4% dos indivíduos deficientes, foi encontrada a variante G6PD Mediterrânea, mutação 563(C-T) e atividade enzimática média de 0,25 UI.g de Hb-1.min-1 a 37ºC, correspondendo a 2,1% da atividade enzimática da G6PD B. A utilização das técnicas tradicionais, aliadas à identificação da variante molecular, são importantes na compreensão das propriedades estruturais, funcionais e comportamento hemolítico dos glóbulos vermelhos do paciente...

Deficiencia de glucosa 6 fosfato deshidrogenasa en niños: caso clínico / Glucose-6-phosphate dehydrogenase deficiency in children: a case report

Introduction: Glucose-6-phosphate dehydrogenase deficiency (G6PD deficiency) is the most common red blood cell (RBC) enzyme disorder. The decrease as well as the absence of the enzyme increase RBC vulnerability to oxidative stress caused by exposure to certain medications or intake of fava beans. Among the most common clinical manifestations of this condition, acute hemolysis, chronic hemolysis, neonatal hyperbilirubinemia, and an asymptomatic form are observed. Objective: To analyze the case of a child who presented hemolytic crisis due to favism. Case report: A 2 year and 7 month old boy with a history of hyperbilirubinemia during the newborn period with no apparent cause, no family history of hemolytic anemia or parental consanguinity. He presented a prolonged neonatal jaundice and severe anemia requiring RBC transfusion. An intake of fava beans 48 h prior to onset of symptoms was reported. G6PD qualitative determination was compatible with this enzyme deficiency. Conclusion: G6PD deficiency can be highly variable in its clinical presentation, so it is necessary to keep it in mind during the diagnosis of hemolytic anemia at any age.

Introducción: La deficiencia de la glucosa 6-fosfato deshidrogenasa (G6PD) es el trastorno enzimático más frecuente del glóbulo rojo (GR). Tanto la disminución como la ausencia de la enzima aumentan la vulnerabilidad del GR al estrés oxidativo provocado por algunos fármacos o la ingesta de habas. Sus manifestaciones clínicas más frecuentes son hemolisis aguda, hemolisis crónica, hiperbilirrubinemia neonatal, y una forma asintomática. Objetivo: Presentar el caso de un niño que debutó como crisis hemolítica debida a favismo. Caso clínico: Varón 2 años 7 meses con antecedente de hiperbilirrubinemia en el período neonatal sin causa evidente, sin historia familiar de anemia hemolítica ni de consanguinidad paterna. Debutó con un cuadro de ictericia y anemia severa que requirió transfusión de GR. Como antecedente anamnéstico se detectó la ingesta de habas 48 h previo al inicio de los síntomas. La determinación cualitativa de G6PD fue compatible con deficiencia de esta enzima. Conclusión: La deficiencia de G6PD puede ser muy variable en su expresión clínica, por lo cual es necesario tenerla presente dentro del diagnóstico diferencial de las anemias hemolíticas a toda edad.

Triagem neonatal de deficiência de glicose-6-fosfato desidrogenase e prevalência das mutações G202A (G6PD A-) e C563T (G6PD Mediterrâneo) em Mato Grosso/Brasil / Neonatal Screening for glucose-6-phosphate dehydrogenase deficiency and prevalence of G202A (G6PD A-) and C563T (G6PD mediterranean) mutations in Mato Grosso / Brazil

Objetivos: A deficiência de glicose-6-fosfato desidrogenase (G6PD) está associada a um maior risco de encefalopatia bilirrubínica e de crise hemolítica aguda grave desencadeada por drogas como a primaquina e a dapsona. Conhecer a prevalência dessa deficiência enzimática em área onde a malária e a hanseníase ainda estão presentes e conhecer a prevalência das principais mutações traz subsídios para planejamento de estratégias com vistas à redução de riscos associados a esta deficiência enzimática. Métodos: Estudo descritivo transversal conduzido em uma região do centro-oeste do Brasil. Exame de triagem para deficiência de G6PD foi realizado em 3573 recémnascidos. Exame confirmatório foi necessário em 188 crianças triadas como possíveis portadores de deficiência. Nas crianças em que foi confirmada a deficiência de G6PD foi feita pesquisa das mutações G202A (G6PD A-) e C563T (G6PD Mediterrâneo) por PCR. Resultados: A deficiência de G6PD foi confirmada em 63 crianças, sendo 60 meninos (95,2%) e três meninas (4,8%). O percentual de exames falso-positivos na fase de triagem foi de 66,5%, estando o percentual de falso-positivos associado à temperatura e tempo de transporte das amostras. Entre as crianças que confirmaram deficiência de G6PD, foi mais frequente a história de anemia em familiares e de icterícia neonatal. Houve associação entre hematócrito baixo e deficiência enzimática, mas não com hemoglobina, contagem de reticulócitos ou neutrófilos. A prevalência da deficiência de G6PD (IC95%) foi de 1,76% (1,37; 2,24) entre os recém-nascidos triados e de 3,34% entre os meninos (2,58; 4,25). A mutação C563T não foi identificada em nenhuma criança, mas a mutação G202A estava presente em 58 crianças - 92,06% (IC95%: 83,29 - 97,03): 56/60 meninos e em 2/3 meninas homozigotas. Foi identificado um menino com Kernicterus portador da mutação G202A em hemizigose. Conclusão: O elevado percentual de falso-positivos na etapa de triagem, o tempo necessário entre coleta...

Objective: Glucose-6-phosphate dehydrogenase (G6PD) deficiency is associated with an increased risk of bilirubin encephalopathy in neonates and acute hemolytic crisis triggered by drugs such as primaquine and dapsone. In an area where malaria and Hansen's disease are still present, knowing the prevalence of this enzyme defect and determining the prevalence of major mutations is important for planning strategies for reducing the risks associated with this enzyme deficiency. Methods: Sectional study was conducted in a Midwestern region of Brazil. Screening for G6PD deficiency was performed in 3,573 neonates. Confirmatory tests were necessary for 188 positively screened children. After confirmation, PCR investigation was utilized to identify the mutations. Results: G6PD deficiency was confirmed in 63 children: 60 boys (95.2%) and 3 girls (4.8%). The percentage of false-positive cases in the screening phase, 66.5% and was associated with the temperature and transportation time of the samples. Family history of anemia and jaundice was more frequent among the children with confirmed G6PD deficiency. An association between a low hematocrit and enzyme deficiency was observed. However, there was no association with hemoglobin reticulocyte or neutrophils counts. The prevalence of G6PD deficiency (CI95%) was 1.76% (1.37; 2.24) among all screened neonates and 3.34% (2.58; 4.25) among male children. The C563T mutation was not identified in any child. The G202A mutation was present in 58 children - 92.06% (CI95%: 83.29 - 97.03), 56/60 boys and 2/3 homozygous girls. One boy with a hemizygous G202A mutation was identified as having Kernicterus. Conclusion: The high percentage of false-positive results when first screening for G6PD deficiency; the long delay time between the test and result; along with the high cost of the this screening test, are all factors that do not support adding this test to the already established Brazilian neonatal screening programs. The prevalence...

Enzyme kinetics and molecular modeling studies of G6PDMahidol associated with acute hemolytic anemia.

G6PDMahidol enzyme is the most common variant in the Achang Chinese ethnic group and clinically manifests as class II. In this study, G6PDMahidol enzyme was characterized by molecular modeling to understand its kinetics. G6PDMahidol, G6PDG487A and G6PDWT proteins were heterologously expressed in the G6PD-deficient DF213 E. coli strain, purified and their steady-state kinetic parameters were determined. Compared with G6PDWT, the Km and Vmax of NADP+ with G6PDG487A were about 28-fold and 12-fold lower, respectively. The Ki values of dehydroepiandrosterone (DHEA), NADPH and ATP with G6PDG487A showed 29.5-fold, 2.36-fold reduction and 1.83-fold increase, respectively. A molecular modeling of G6PDG487A was performed based on the X-ray structure of human G6PD (PDB: 2BH9). It is suggested that Ser-163 might affect the stability of G6PDG487A -helix d and -strand E, besides the conformation of -strand D. In conclusion, the biochemical and structural properties of G6PDG487A and G6PDWT enzymes are significantly different, which may be responsible for clinical diversity of G6PD deficiencies.

Prevalence of Gilbert syndrome in parents of neonates with pathologic indirect hyperbilirubinemia

The cause of hyperbilirubinemia cannot be found in about 45% of cases of neonatal jaundice. Gilbert syndrome [GS] is the most common congenital disease associated with bilirubin metabolism in the liver. Since the screening value of genetic tests cannot be fully determined until accurate data on the prevalence and penetrance of the GS genotype are known, we sought to estimate whether the prevalence of GS is higher in the parents of neonates with severe unexplained indirect hyperbilirubinemia. Case-control study of parents of neonates with severe unexplained indirect hyperbilirubinemia admitted to a neonatal ward. We used the rifampin test [checked bilirubin before and 4 hours after administration of 600 mg rifampin] for diagnosis of GS in parents of 115 neonates with severe unexplained indirect hyperbilirubinemia. We compared the prevalence of GS in these parents with that of a control group of 115 couples referred for premarital counseling. The 115 neonates were aged 5.2 [1.6] days [mean, standard deviation], all were breast-fed, and males constituted 56.5%. Mean total serum bilirubin [TSB] level was 20.96 [5.48] mg/dL. 14.8% were glucose 6 phosphate dehydrogenase [G6PD] deficiency was present in 14.8%, and 10.4% had A, B or O blood group [ABO] incompatibilities with their mothers. There was no difference in the prevalence of GS between parents of the group with hyperbilirubinemia [22.2%] and the control group [19.13%] [P=.42]. Mean TSB in neonates with parents who had GS was more [about 3 mg/dL] than in neonates with normal parents [P=.004]. Fathers had GS twice as often as the mothers among the parents of neonates with hyperbilirubinemia [P=.003], among the control group [P=.009] and among neonates [P=.014]. This study showed that GS cannot cause severe indirect hyperbilirubinemia by itself, but it may have a summative effect on rising bilirubin when combined with other factors, for example, G6PD. Our results showed that in GS, males are affected about twice as much as the females

Brucellosis triggering hemolytic anemia in glucose-6-phosphate dehydrogenase deficiency

To present a case of acute brucellosis triggering acute hemolytic anemia in a subject with glucose-6-phosphate dehydrogenase [G6PD] deficiency. A 17-year-old male patient presented with fever, malaise and jaundice. His blood and bone marrow cultures yielded Brucella species. In addition, he was found to have acute hemolytic anemia due to previously undiagnosed G6PD deficiency. He was started on folic acid supplementation and given a combination of doxycycline and rifampicin for 6 weeks. His response to antibiotic therapy was optimal; the hemolytic anemia resolved. There were no further episodes of hemolysis. This case showed that the differential diagnosis of acute hemolytic anemia in subjects with G6PD deficiency should include brucellosis, especially in regions where the infection is endemic

hemolysis trigger in glucose-6-phosphate dehydrogenase enzyme deficiency vicia sativa [Vetch]

Glucose-6-phosphate dehydrogenase [G6PD] is an enzyme, playing an important role in the redox metabolism of all aerobic cells. It was reported that certain medications, fava beans, and infections can trigger acute hemolytic anemia in patients with G6PD deficiency. An 8-year-old male patient was admitted to the hospital with blood in the urine, headache, dizziness, fatigue, loss of appetite, and jaundice in the eyes, 24 hours after eating large amounts of fresh, vetch grains. Laboratory investigation revealed hemolytic anemia, hyperbilirubinemia, and G6PD deficiency. Approximately 0.5% of fava bean seeds have 2 pyrimidine beta-glycosides called, vicine and convicine. Vetch has 0.731% vicine, 0.081% convicine, and 0.530% beta cyanoalanine glycosides. The aim of this case report is to emphasize the importance of vetch seeds as a cause for hemolytic crisis in our country, where approximately one million tons of vetch is produced per year, especially in the agricultural regions

Associação de metemoglobinemia e deficiência de glicose-6-fosfato desidrogenase em pacientes com malária tratados com primaquina / Association of methemoglobinemia and glucose-6-phosphate dehydrogenase deficiency in malaria patients treated with primaquine

Este estudo teve como objetivo verificar a ocorrência de metemoglobinemia em indivíduos deficientes da glicose-6-fosfato desidrogenase durante o tratamento da infecção malárica com primaquina. Foram selecionados pacientes com diagnóstico para malária por Plasmodium vivax ou mista V+F (Plasmodium vivax + Plasmodium falciparum), Grupo 1: com 74 indivíduos com diagnóstico clínico de metemoglobinemia e Grupo 2: 161 indivíduos sem diagnóstico clínico de metemoglobinemia. Quanto à deficiência da G6PD, nos Grupos 1 e 2, houveram 51,3 por cento (38) e 8,7 por cento (14) de indivíduos enzimopênicos, respectivamente, demonstrando através de tais dados, significância estatística na associação com a metemoglobinemia somente nos indivíduos do Grupo 1 (p<0,05). A comparação da relação da metemoglobinemia à deficiência da G6PD mostrou haver uma possível associação de indivíduos enzimopênicos desenvolverem metemoglobinemia com maior freqüência.

This study had the aim of investigating occurrences of methemoglobinemia among individuals with glucose-6-phosphate dehydrogenase deficiency during treatment for malaria infection using primaquine. Patients with a diagnosis of malaria caused by Plasmodium vivax or the V+F mixture (Plasmodium vivax + Plasmodium falciparum) were selected. Group 1 consisted of 74 individuals with a clinical diagnosis of methemoglobinemia and Group 2 consisted of 161 individuals without a clinical diagnosis of methemoglobinemia. The glucose-6-phosphate dehydrogenase deficiency rates (numbers of enzymopenic individuals) in Groups 1 and 2 were 51.3 percent (38) and 8.7 percent (14) respectively. These data demonstrated a statistically significant association with methemoglobinemia only among the individuals in Group 1 (p<0.05). Investigation of the relationship between methemoglobinemia and glucose-6-phosphate dehydrogenase deficiency showed that there was a possible association such that enzymopenic individuals may develop methemoglobinemia more frequently.

Acute Hemolysis and Renal Failure caused by Hepatitis A Infection with Underlying Glucose-6-Phosphate Dehydrogenase Deficiency / 대한진단검사의학회지

Although glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common enzyme disorder worldwide, it has rarely been reported among Korean. The patient was previously healthy 39 yr old male who showed severe hemolytic anemia and acute renal failure accompanied by hyperbilirubinemia after hepatitis A infection. The additional studies for differential diagnosis of hemolytic anemia showed a moderate deficiency of G6PD enzyme. Because hepatitis A infection in patient with G6PD deficiency present much more severe clinical symptoms, G6PD enzyme should be examined in patients with triggering factors of hemolysis such as hepatitis A infection.

Neonatal screening for glucose-6-phosphate dehydrogenase deficiency.

OBJECTIVE: This study was carried out to detect the incidence of erythrocytic Glucose-6 -Phosphate dehydrogenase (G-6-PD) deficiency, to compare the incidence of hyperbilirubinemia in G-6-PD deficient neonates as compared to G-6-PD normal neonates and to asses the usefulness of neonatal screening for G-6-PD deficiency. METHOD: In a retrospective hospital based study 2,479 male and female neonates consecutively born at Indraprastha Apollo hospital between July 1998 to June 2003 who were screened for G-6-PD levels were evaluated for the incidence of G-6-PD deficiency. RESULTS: Incidence of G-6-PD deficiency was found to be 2.0%. Incidence in males was 283% and female was 1.05%. The incidence of hyperbilirubinemia was found to be 32% in G-6-PD deficient neonates which was significantly higher than the incidence of hyperbilirubinemia in neonates with normal G-6-PD, which was 12.3% (P< 0.001). CONCLUSION: Our data suggests that neonatal screening for G-6-PD deficiency is a useful test for preventing and early treatment of complications associated with it.

G 6 PD deficiency induced hemolysis in diabetic twins.

We report hemolysis due to glucose-6-phosphate dehydrogenase (G-6-PD) deficiency in two Egyptian monozygotic brothers, at the onset of diabetes type 1. Hemolysis occurred following the treatment of hyperglycemia and ketoacidosis (one twin). It was related to unknown G-6-PD deficiency. The fall in glucose availability after the treatment of hyper glycemia is proposed as a possible etiology for hemolysis.

Glucose-6-phosphate dehydrogenase deficiency in India.

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the commonest red cell enzymopathy in humans and has an X-linked inheritance. It has been reported from India more than 30 years ago and the prevalence varies from 0-27% in different caste, ethnic and linguistic groups. The major clinical manifestations are drug induced hemolytic anemia, neonatal jaundice and chronic non-spherocytic hemolytic anemia. Individuals with G6PD deficiency have a selective advantage against falciparum malaria. Thirteen biochemically characterized variants have been reported from India. At the molecular level, G6PD Mediterranean is the most common deficient variant in the caste groups whereas, G6PD Orissa is more prevalent among the tribal of India. The third common variant seen in India is G6PD Kerala-Kalyan.

Glucose-6-phosphate dehydrogenase deficiency in neonates.

OBJECTIVE: Glucose-6-phosphate dehydrogenase (G6PD) deficiency is an inherited deficiency that may be the cause of neonatal hyperbilirubinemia, as has been found in several countries and among widely different ethnic groups, especially in Mediterranean region. Our aim was to study the prevalence of G6PD deficiency in relation to neonatal jaundice. METHODS: From March 1998 to April 2001 we studied 705 clinically icteric neonates who were admitted to Al-Zahra and Beheshti hospitals, two teaching hospitals in Isfahan, Iran. Laboratory investigations included determination of direct and indirect serum bilirubin concentrations, blood group typing, direct coomb's test, hemoglobin, blood smear, reticulocyte count and G6PD level. RESULTS: In only 53 (7.5%) of cases G6PD deficiency was diagnosed. In all G6PD deficient neonates no evidence of other factors known to cause hyperbilirubinemia were detected. The sex distribution was 13 (24.5%) females and 40 (75.5%) males in the G6PD deficient group. The mean bilirubin level in G6PD deficient and G6PD normal groups were 22.26 +/- 8.36 and 18.14 +/- 3.85 mg/dl, respectively (p=0.001). Phototherapy was required in G6PD deficient and other icteric neonates with duration of 3.76 +/- 1.93 and 3.13 +/- 2.14 days, respectively (p=0.045). Twenty-seven of the 53 (50.9%) G6PD deficient infants required exchange transfusion. None of them developed kernicterus. CONCLUSIONS: Since the prevalence of severe hyperbilirubinemia among our neonates was relatively high and about half of them required exchange transfusion, early detection of this enzymopathy regardless of sex and close surveillance of the affected newborns may be important in reducing the risk of severe hyperbilirubinemia and exchange transfusion.

Glucose-6-phosphate dehydrogenase deficiency in Thailand; its significance in the newborn.

The prevalence of G6PD deficiency in Thai males ranges from 3-18% depending upon the geographic region. G6PD "Mahidol" (163 Gly --> Ser) is the most common variant found in the Thai population. Almost all affected Thai individuals are not anemic and are asymptomatic. Severe acute intravascular hemolysis is occasionally seen, for instance, in those cases who have a viral infection, bacterial infection or have been exposed to chemicals or drugs. In Thailand, diagnosis of G6PD deficiency is usually made only in symptomatic cases. Neonatal screening of G6PD deficiency is not practiced nationwide, though studies have been done in several institutes. The assessment of G6PD activity in the newborn is mostly in order to find out the cause of neonatal jaundice. In our experience and that of others. G6PD deficient newborns are more prone to develop neonatal jaundice which is, on its own, no more severe than jaundice from other causes. Kernicterus due to G6PD deficiency, though still seen, is now very rare. Awareness of the hazard of hyperbilirubinemia, whatever the cause, along with active management is needed to prevent the occurrence of kernicterus. Neonatal screening is useful to detect abnormalities in the newborn. Weighing of the cost and benefit of neonatal screening should be made and the families of patients should be offered proper education and counseling to help them understand their babies' condition.

Mass newborn screening for glucose-6-phosphate dehydrogenase deficiency in Singapore.

Recognition of kernicterus as a significant contributor to newborn mortality and morbidity in G6PD deficient babies and the ease with which G6PD deficiency could be detected and kernicterus prevented by avoidance of triggers, led to the establishment of mass newborn screening for G6PD deficiency in 1965. G6PD deficient newborns are identified within a day of birth by measuring the enzyme activity in cord blood. They are then physically protected from triggers by keeping them in the hospital for the first 2 weeks of life after their parents are counseled. Enzyme activity is measured using Wong's in-house modification of the Bernstein's technique or the BM G6PD Deficiency Screening test based on the Beutler assay. Close to 1.6 million newborns, representing practically 100% of all births have been screened. Analysis of data from 22,830 newborns at the National University Hospital reveals an incidence of 1.62% in all newborns. 3.15% in males and and 0.11% in females. A distinct racial variation in the incidence of deficiency was observed in males: Chinese 3.94%, Malays 2.95% and Indians 0.66%. Intermediate deficiency was most frequently identified (1.83%) in Chinese females. With the preventive measures. the incidence of kernicterus has dropped dramatically and there has been, over the last 20 years, no reported cases of kernicterus in newborns with G6PD deficiency. We are now looking at issues like comparing different assay techniques and determining a shorter period of stay in hospital. We believe that all Asians, especially Chinese babies, should be screened for G6PD deficiency, irrespective of which country the child is born.

A simple, rapid fluorometric assay for the determination of glucose 6-phosphate dehydrogenase activity in dried blood spot specimens.

Glucose 6-phosphate dehydrogenase (G6PD) deficiency is the most common enzymopathy of human beings and is the most common cause of jaundice and acute hemolytic anemia in South East Asia. The deficiency causes acute hemolytic anemia following ingestion of 6-amino quinoline antimalarials, phenacetin, and other substances. The rapid identification of infants or patients with this deficiency would help to prevent their exposure to these substances and subsequent risk to health. The assay is relatively simple. A 3mm punch from a dried blood spot sample is placed in a well of a black fluorescent microtiter plate containing calibrators and controls in duplicate. 100 microl of reagent is added and the sample is allowed to react for 30 minutes at ambient temperature after which 200 microl of stop reagent is added. The plate may be read immediately or up to one hour in a fluorescent reader (ex 355 nm: em 460 nm). Glutathione. ascorbate and bilirubin do not affect the assay. hemoglobin does quench the fluorescence by about 1.1 fluorescence units/g/dHb. This would not cause any false negatives and deficients would not be missed. G6PD activity in whole blood normal samples was examined at -20, 6 and 37 degrees C over 14 days. The samples lost about 20% activity after 48 hours and 31% by the end of 14 days. The samples stored at -20 degrees C and 6 degrees C remained relatively stable over this period. In a preliminary study eight diagnosed G6PD deficient samples had a mean value of 2.0 U/gHb (range 0.8 to 4.4) and fell within 3 SD units of the mean. Forty one normal samples had a mean of 6.6 micromol/min/gHb. Only one sample with a low hemoglobin level fell outside of 3 SD units of the mean. The Wallac assay was compared to the Sigma G6PD assay and although the values appeared lower at normal levels, the deficient samples compared well.