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Objective:To investigate the diagnostic value of fluorescence quantitative method and G6PD/6PGD ratio method in glucose-6-phosphate dehydrogenase (G6PD) deficiency and the type of gene mutation.Methods:A total of 1 201 patients (711 males and 490 females) with suspected G6PD deficiency in Shanghai Children′s Hospital were collected from June 2018 to March 2021. Fluorescence quantification method, G6PD/6PGD ratio method and multicolor melting curve were used to detects enzyme activity, ratio and gene mutation type. Comparison of each index and evaluation of its diagnostic efficiency were performed.Results:Among 1 201 suspicious samples, 163 cases (135 males and 28 females) were finally diagnosed. 156 cases were diagnosed by fluorescence quantitative method with a detection rate of 95.71%, and 140 cases were diagnosed by G6PD/6PGD ratio method with a detection rate of 85.89%. enzymatic activity of G6PD and ratio of G6PD/6PGD in male were significantly lower than female, and the differences were statistically significant ( U=642.5, 734.5, P<0.001). 112 cases received G6PD gene mutation detection and 92 cases were diagnosed, 74 were hemizygous mutations, 1 were homozygous mutations, 15 were heterozygous mutations, and 2 were compound heterozygous mutations. Among 15 cases of heterozygous mutations, 11 cases were diagnosed by fluorescence quantitative method, the diagnosed rate was 73.33%, 4 cases were diagnosed by G6PD/6PGD ratio method, and the diagnosed rate was 26.67%. A total of 7 mutation sites were detected and the proportions were c.1388G>A (32.22%), c.1376G>T (30.00%), c.871G>A (13.33%), c.1024C>T (11.11%). c.95A>G (7.78%), c.487G>A (4.44%), c.392G>T (1.11%). The enzymatic activities of c.1376G>T and c.1024C>T, c.487G>A were statistically significant ( P<0.001,0.015); the G6PD/6PGD ratios of c.1024C>T and c.1388G>A, c.1376G>T were statistically significant ( P=0.017,0.002,0.011,0.013). Fluorescence quantitative method had sensitivity of 100%, specificity of 95.65%, and the area under the curve (AUC) is 0.972. The sensitivity of the G6PD/6PGD ratio method was 100%, the specificity was 94.57%, and the AUC was 0.979. The sensitivity of fluorescence quantitative method combined with G6PD/6PGD ratio was 96.7%, the specificity was 100%, and the AUC was 0.992. Conclusions:Compared with fluorescence quantification, the G6PD/6PGD ratio method might not be able to diagnose female heterozygotes effectively; The panel of G6PD fluorescence quantification and G6PD/6PGD ratio was helpful to reduce the missed diagnosis. Combined with gene mutation analysis, it could improve the diagnosis rate of G6PD deficiency in the children.
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Objective To evaluate the feasibility of PerkinElmer Genetic Screening Processor(GSP) in the application of newborn screening for congenital hypothyroidism (CH) and congenital adrenal hyperplasia (CAH) by detecting thyroid-stimulating hormone (TSH) and 17-OH-progesterone(17-OHP).Methods The dried-blood spots specimens from Centers for Disease Control and Prevention(CDC) and the quality control in the reagent kit were detected and the accuracy,precision and linearity were calculated.A total of 1 012 samples of TSH(60 of positive and 952 negative samples) and 991 samples of 17-0HP(34 positive and 957 negative samples)were detected.The initial cut-off value was determined by ROC curve determined.The consistency between the results from GSP and clinical diagnosis was analyzed.Results The average of within-run coefficient of variation(CV) of TSH and 17-OHP were 6.69％ to 12.6％ and 7.52％ to 9.29％,and the average of between-run CV were 6.91％ to 10.96％ and 6.86％ to 12.36％,respectively.The average of bias of TSH and 170HP were-14.28％ to-0.74％ and-0.45％ to 12.54％.The linearity of GSP detection was fine.The initial cut-off values were 23.43 U/mL(TSH) and 21.42 ng/mL(17-OHP).The sensitivity of GSP detection was 100％ and the specificity of TSH and 17-OHP were 98.11％ and 99.58 ％ respectively.The results of GSP detection showed good consistency with clinical diagnosis.Conclusion As the first real automatic fluorescence immunoassay analyzer,GSP could be used in routine clinical diagnosis for CH and CAH.
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Objective To summarize the acylcarnitine profile in children with malnutrition,with an attempt to distinguish it from those of medium-chain acyl-CoA dehydrogenase (MCAD) deficiency,multiple acylCoA dehydrogenase (MAD) deficiency,or glutaric aciduria type Ⅱ (GA Ⅱ).Methods Thirteen pediatric patients with malnutrition and 214 children of the same age but without malnutrition,which was set as the control group,were included in this study.The blood samples were collected at admission,and the concentration of carnitine and acylcarnitines were measured in bloodspots by tandem mass spectrometry using samples nnderivatized.Results The concentrations of acylcarnitines which were involved in fatty acid oxidation,including octadecanoyl (C18) to acetyl (C2) acylcarnitines and ketonic acylcarnitines,were higher in malnutrition group than in the control group.Particularly,the concentration of decanoyl acylcarnitine (C10) in the malnutrition group was (0.203 ±0.105) μmol/L,which was out of the normal rang (0-0.200 μmol/L),was significantly higher than that [(0.054 ±0.030) μmol/L] in the control group (P ＜0.001).There was no significant difference in the concentrations of acylcarnitines [e.g.propionyl (C3),isovaleryl (C5),3-hydroxy-isovaleryl (C5OH),and glutaryl (C5DC) acylcamitines] involved in amino acid decomposition between the malnutrition and control groups.Conclusions The concentrations of acylcarnitines related to fatty acid oxidation elevate in children with malnutrition.In particular,the medium-chain acylcarnitines C10 is out of the normal range,which can be used to differentiate malnutrition from MCAD and MAD.
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Objective To provide methodological references for laboratories to carry out newborn screening for disorders of amino acid metabolism,we compared the difference and distribution of ten amino acids including alanine (Ala),arginine (Arg),citrulline (Cit),glycine (Gly),leucine (Leu),methionine (Met),ornithine (Orn),phenylalanine (Phe),tyrosine (Tyr),and valine (Val) from newborn dried blood spots specimen using derivatization or non-derivatization as sample preparation methods.Method It is a comparative research study.A total of 4135 newborn screening dried blood spot samples for inborn errors of metabolism were collected from January to June,2012.All specimens came from neonatal screening center of shanghai children's hospital.Samples were prepared by two different techniques,the corresponding kits and the procedures were used as follows:(1) Simultaneous detection of 100 dried blood spot specimens using two methods respectively to compare the paired difference of each amino acid.(2) 2000 cases of normal newborn specimens were detected respectively to obtain the normal distribution of ten neonatal amino acids.(3) 35 specimens from patients previously diagnosed positively as inborn errors of metabolism were simultaneously detected with 7 amino acids to verify the consistency of two techniques in clinical judgment.Results The amino acid levels of normal newborns analyzed by one-sample.kolmogorovSmirnov test (Z value ranged from 1.997 to 6.229) showed a skewed distribution (P ＜ 0.01).Except for Leu and Tyr,non-derivatization techniqueshowed a lower concentration than derivatization technique,and the CVs of nine amino acids were ＜ 10％ except for Met (the CV of Met was 47.8％),and the average CV is 7.8％.Except for Met,Phe and Tyr,the levels of other 7 amino acids measured by two techniques showed no significant difference (P ＞ 0.05).According to 0.5th to 99.5th percentiles,the normal reference range for derivatization method were greater than on-derivatization method,and the average value was 25.3％.After clinic judgment,the results of the abnormal indicators of children with true metabolism disorders showed no statistically significant between two methods (P ＞ 0.05),the detection rate was 100％.Conclusions There was a slight difference between derivatization and the non-derivatization techniques in detecting multiple amino acids.The results of the abnormal indicators of amino acid metabolism disordersshow no statically significant difference between the two methods,and no difference in clinical judgment.Both methods can be used in detecting amino acid metabolism disorders in newborn screening.
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Objective To develop a method of detecting dry blood spot (DBS) samples collected in the filter paper by using Surface Enhanced Raman Spectroscopy (SERSp,also known as Raman molecular fingerprint spectrum) technology.The possibility and reliability of applying the technology to the newborn screening of congenital hypothyroidism (CH) was investigated.Methods The case-control study was used.The results of CH-hTSH screening test based on the DBS samples collected in the filter paper,and the clinical diagnosis were all provided by neonatal screening center of Shanghai Children's Hospital.The results of SERSp analysis were provided by Dalian University of Technology.Six positive and six negative samples,which were confirmed respectively by CH-hTSH screening using time resolved fluoroimmunoassay (TRFIA) were chosen.With the spectrum lines of 725 cm-1 as internal standard,the SERSp from the aqueous solution of these samples were analyzed.The specific spectrum line analysis,the principal component analysis (PCA) and the scatter diagram of PCA were used to compare the results of TSH test.Results With the spectrum lines of 725 cm-1 as internal standard,the CH specific spectrum lines of 1373 cm-1 and 1400 cm-1 were discovered.They showed the most significant difference of the peak height between the positive and negative samples of CH,while TSH specific spectrum lines of 785 cm-1,827 cm-1 and 853 cm-1 failed to distinguish positive from negative samples with distinct peak height.The consistency between the results of CH-Raman Screening and CH-hTSH Screening was 91.7％ (11/12),which was proven by PCA,the scatter diagram of PCA and the specific spectrum line analysis.The sensitivity of CH-Raman Screening was 100％ and the specificity was 83.3％.Conclusion There is a new CH-Raman Screening method using the SERSp analysis,which was proven to be a promising technology in the newborn screening of congenital hypothyroidism with the DBS in the filter paper.
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ObjectiveTo evaluate the influence of electrotome on permanent and temporary cardiac pacemaker in laparoscopic cholecystectomy ( LC),and the application of cardiac pacemaker to the cases of cholecystolithiasis combined with bradyarrhythmia.MethodsClinical data of 215 patients with permanent or temporary cardiac pacemaker who underwent were studied for the preoperative and postoperative variation of pacemaker function,and for the influence of electricity coagulation during the operation on cardiac pacemaker function.ResultsLC was successfully completed in all 215 patients.The function of cardiac pacemaker was not obviously interfered during the operation,and the parameters of cardiac pacemaker did not remarkably change after the operation.ConclusionCardiac pacemaker is slightly interfered when electrotome and electrocoagulation were used in LC; LC is feasible and safe for patients with bradyarrhythmia by placement of cardiac pacemaker.
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Objective To investigate the blood levels of amino acids in children aged 0-15 year, with an attempt to provide evidence for evaluating amino acid status and diagnosing metabolic diseases of amino acid.Methods The blood levels of eleven amino acids in 1900 children aged 0-15 years were determined by tandem mass spectrometry (MS/MS). Results The blood levels of leucine & isoleucine, valine, phenylalanine tyrosine,glycine, proline, ornithine, and alanine gradually decreased after birth, reaching the lowest levels at the ages of 4-6 months, and then gradually increased, reaching the normal range at the ages of 7 months-1 year. The blood levels of alanine and glycine reach the second peaks on the ages of 9 years in girls and on 11 years in boys. The blood levels of methionine and arginine were lowest in the first week of age, became highest in 1-3 months, decreased to the normal ranges after 4-6 months, and kept the level afterwards. The ratios between prosomatic amino acid and productive amino acid, between ornithine and arginine, between citrulline and arginine, and between ornithin and citrulline were highest in the first week of age and decreased to normal values after 3 to 12 months. The concentrations of amino acids in group of 7 months-15 years were significantly different from the group of 1 day-1 month and group of 2-6 months (P ＜0. 05 or P ＜0. 01 ). The concentrations of amino acids were significantly higher in females than in males in the group of 1 day-1 month and in all age groups (P ＜0. 05 or P ＜0. 01 ) . Conclusions The concentrations and profiles of amino acids change remarkably during the first year of age. Age should be carefully considered when evaluating the nutritional status of amino acid and diagnosing metabolic diseases of amino acids.
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Objective To develop the newborn screening and treatment for congenital hypothyroidism (CH) in order to prevent the children from growth and mental retardation and to improve the population quality in China. Methods TSH value in the dried-blood spot speciments on the filter papter was measured by time-resolved fluorescence immunoassay (Tr-FIA). Clinical analysis for the CH patients was performed. Results Among 315 472 neonates screened for CH, 66 (47 overt and 19 subclinical) cases of CH were confirmed and the incidence of CH was 1/4 780. Among the 47 typical cases, 2 infants were recognized with transient hypothyroidism, 2 infants were not reevaluated because of below 2 year old, 1 infant died, 1 case gave up treatment, the rest 41 infants were permanent CH. Among the 19 atypical cases, 7 cases were recognized with permanent CH, 4 cases with transient disease, 5 cases with hyperthyrotropinemia, 3 cases were not evaluated because of below 2 years old at that time. The patients followed in the clinic are all currently with normal growth and intellectual development. Conclusion Our center is the first labolatory to screen CH with Tr-FIA in China. Tr-FIA is an ideal non-radioactive technology. The typical patients as well as atypical cases should be all treated using L-thyroxine. It is essential to arrange a trial treatment for confirming the diagnostic results from screening and to decide whether they need a lifetime treatment.
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Objective: To study the dietary habits and serum leptin levels of obese children and its possible association to lipids and glucose metabolism, serum lipids and C peptide (CP) in children with different body weight. Methods: 52 obese children were inquired about their favorite foods and food intake. 145 healthy children (85 boys, 60 girls), aged 6-13, with a wide range of body mass index (BMI) 14 to 30 kg/m 2, were measured for fasting serum leptin, CP, glucose, lipids (TG, TC, LDL C,HDL C), height and weight. The children were classified into three groups by BMI, i.e. normal weight group (A group), overweight and slight obese group (B group). moderate and severe obese group (C group). Results: 65% of obese children liked meat best. 38.46% of them ate most in supper. The serum leptin, CP, TG in obese children were higher than that in normal children. Leptin concentration of girls were higher than that in boys in A and B groups (P