Sequence variants in the BTD underlying biotinidase deficiency in families of Pakistani origin.

BACKGROUND: Biotinidase deficiency (BTD) is a rare autosomal recessive metabolic disease, which develops neurological symptoms because of the impaired biotin recycling. Pathogenic mutations on BTD gene cause BTD deficiency. The clinical features and mutation analysis of Pakistani children with BTD deficiency have rarely been described. Herein, for the first time, we report the clinical features, BTD gene mutations and biochemical analysis of seven symptomatic children with BTD deficiency from Pakistan. METHODS: Seven suspected BTD-deficient patients who presented abnormal organic acid profiles and clinical features were subjected to Sanger sequencing to identify pathogenic mutations in the BTD gene. The results were analyzed by Mutation Surveyor Software. RESULTS: All seven patients exhibited common biotinidase deficiency symptoms including hypotonia, developmental delay and seizures. Biochemical analysis shows marked excretion of 3-hydroxy isovalerate in all cases, followed by 3-hydroxy propionate and methyl citrate. Sanger sequencing revealed one frame-shift mutation, c.98_104delinsTCC (p.Cys33Phefs), and two missense mutations, c.1612C>A (p.Arg538Ser) and c.1330G>C (p.Asp444His). All mutations were in the homozygous state and classified as pathogenic in published studies and mutation databases. CONCLUSIONS: This study has validated the BTD variants as the underlying cause of biotinidase deficiency in which molecular testing of BTD is supported by urinary organic acid analysis and clinical diagnosis. Secondly, the strength of the local availability of this test in Pakistan will paved the way for the neonatal screening of biotinidase deficiency.

Insights from semi-structured interviews on integrating artificial intelligence in clinical chemistry laboratory practices.

BACKGROUND: Artificial intelligence (AI) is gradually transforming the practises of healthcare providers. Over the last two decades, the advent of AI into numerous aspects of pathology has opened transformative possibilities in how we practise laboratory medicine. Objectives of this study were to explore how AI could impact the clinical practices of professionals working in Clinical Chemistry laboratories, while also identifying effective strategies in medical education to facilitate the required changes. METHODS: From March to August 2022, an exploratory qualitative study was conducted at the Section of Clinical Chemistry, Department of Pathology and Laboratory Medicine, Aga Khan University, Karachi, Pakistan, in collaboration with Keele University, Newcastle, United Kingdom. Semi-structured interviews were conducted to collect information from diverse group of professionals working in Clinical Chemistry laboratories. All interviews were audio recorded and transcribed verbatim. They were asked what changes AI would involve in the laboratory, what resources would be necessary, and how medical education would assist them in adapting to the change. A content analysis was conducted, resulting in the development of codes and themes based on the analyzed data. RESULTS: The interviews were analysed to identify three primary themes: perspectives and considerations for AI adoption, educational and curriculum adjustments, and implementation techniques. Although the use of diagnostic algorithms is currently limited in Pakistani Clinical Chemistry laboratories, the application of AI is expanding. All thirteen participants stated their reasons for being hesitant to use AI. Participants stressed the importance of critical aspects for effective AI deployment, the need of a collaborative integrative approach, and the need for constant horizon scanning to keep up with AI developments. CONCLUSIONS: Three primary themes related to AI adoption were identified: perspectives and considerations, educational and curriculum adjustments, and implementation techniques. The study's findings give a sound foundation for making suggestions to clinical laboratories, scientific bodies, and national and international Clinical Chemistry and laboratory medicine organisations on how to manage pathologists' shifting practises because of AI.

Inappropriate supplementation of Vitamin D can result in toxicity: a crosssectional study of paediatrics population.

OBJECTIVE: To evaluate children with suspected or definite hypervitaminosis D with respect to prevalence, clinical manifestations and pharmacological aspects. METHODS: The retrospective cross-sectional study was conducted at the Aga Khan University Hospital, Karachi, and comprised medical records from January 1 to December 31, 2018, of children aged <18 years with 25-hydroxyvitamin D levels >50ng/ml. Clinical and pharmacological data was retrieved. Data was analysed using SPSS 23. RESULTS: Of the 118,149 subjects visiting the clinical laboratory during the study period, children tested for serum 25-hydroxyvitamin D levels were 16,316(13.8%) who had a median age of 9.78 years (interquartile range: 10.2 years). Children who registered for consultation were 2720(16.6%), and, out of them, 602(22%) had serum 25-hydroxyvitamin D >50ng/ml. The median 25-hydroxyvitamin D levels and age were 70.1ng/ml (interquartile range: 100ng/ml) and 3.1 years (interquartile range: 17.93 years), respectively, and 345(57.3%) of them were boys. Children supplemented with vitamin D were 197(33.1%) and 193(97.9%) of them were prescribed by physicians. Mega-doses were taken by 68(34.17%), while the remaining had used various combinations in syrup or tablet forms. Commonly prescribed mega-doses were 600,000IU 30((44.1%) and 200,000IU 31(45.5%) injections of vitamin D. The primary indications were pains/aches in 51(25.8%) cases, developmental delay 50(25.3%), and vitamin D deficiency 49(24.8%). The main symptoms of hypervitaminosis D or toxicity were abdominal pain 27(13.7%) and constipation 31(15.7%). CONCLUSIONS: Children should be given vitamin D supplements with caution as prolonged supplementation and repeated mega-doses can result in toxicity which may cause serious consequences.

Factors Associated With Awareness Of Literate Mothers About Newborn Screening: A Cross-Sectional Study From A Low-Middle-Income Country.

Objectives: To explore the awarenesslevel of literate mothersregarding newborn screening programmes, and to evaluate the associated factors. METHODS: The descriptive, cross-sectional study was conducted at the Section of Chemical Pathology, Department of Pathology andLaboratoryMedicine,AgaKhanUniversity,Karachi,fromJanuary toSeptember 2021, andcomprisedmothers aged 18 years or more. Data was collected using a structured questionnaire about newborn screening, and the subjects were compared in terms of age, residential background, education and parity. Data was analysed using SPSS 23. RESULTS: Of the 1016 responses, 896(88.2%) were analysed. The mean age of the sample was 37.7±10.87 years. There were 470(52.4%) mothers aged 31-45 years, 859(95.87%) were from urban areas, 751(84%) had a graduate degree, 652(72.7%) weremultiparous andhad824(91.9%)hadhealthy children.Overall, 386 (43%)mothershadawarenessofnewbornscreening programmes. The main factors associated with awareness were age, education, primiparity, having healthy children, and province ofresidencebeing Sindh andPunjab(p<0.05),while the urban-ruraldividedwas not a significantfactor(p=0.737). Cost of healthcare 417(46.5%) and lack of awareness among physicians 356(39.7%) were identified asthe main challenges in establishing newborn screening servicesin the country. CONCLUSIONS: The awareness among mothers about new born screening programmes was generally low among the subjects studied.

Urinary metabolomics using gas chromatography-mass spectrometry: potential biomarkers for autism spectrum disorder.

BACKGROUND: Diagnosis of autism spectrum disorder (ASD) is generally made phenotypically and the hunt for ASD-biomarkers continues. The purpose of this study was to compare urine organic acids profiles of ASD versus typically developing (TD) children to identify potential biomarkers for diagnosis and exploration of ASD etiology. METHODS: This case control study was performed in the Department of Pathology and Laboratory Medicine in collaboration with the Department of Pediatrics and Child Health, Aga Khan University, Pakistan. Midstream urine was collected in the first half of the day time before noon from the children with ASD diagnosed by a pediatric neurologist based on DSM-5 criteria and TD healthy controls from August 2019 to June 2021. The urine organic acids were analyzed by Gas Chromatography-Mass Spectrometry. To identify potential biomarkers for ASD canonical linear discriminant analysis was carried out for the organic acids, quantified in comparison to an internal standard. RESULTS: A total of 85 subjects were enrolled in the current study. The mean age of the ASD (n = 65) and TD groups (n = 20) was 4.5 ± 2.3 and 6.4 ± 2.2 years respectively with 72.3% males in the ASD group and 50% males in the TD group. Parental consanguinity was 47.7 and 30% in ASD and TD groups, respectively. The common clinical signs noted in children with ASD were developmental delay (70.8%), delayed language skills (66.2%), and inability to articulate sentences (56.9%). Discriminant analysis showed that 3-hydroxyisovalericc, homovanillic acid, adipic acid, suberic acid, and indole acetic were significantly different between ASD and TD groups. The biochemical classification results reveal that 88.2% of cases were classified correctly into ASD& TD groups based on the urine organic acid profiles. CONCLUSION: 3-hydroxy isovaleric acid, homovanillic acid, adipic acid, suberic acid, and indole acetic were good discriminators between the two groups. The discovered potential biomarkers could be valuable for future research in children with ASD.

Development and validation of standard and real patient gallstone library using Fourier transform infra-red spectroscopy.

BACKGROUND: Analysis of the constituents of gallstones using various spectroscopic techniques assists in identification of the pathogenesis of gallstones. In the current study, using Fourier Transform Infra-Red (FTIR) Spectroscopy, a Gallstone Standard Library (GSL) and a Gallstone Real Patients' Library (GRPL) were developed and validated for gallstone composition analysis. METHODS: The study was conducted at the Department of Pathology & Laboratory Medicine, Aga Khan University, Pakistan. Pure standards (cholesterol, calcium carbonate, bilirubin and bile salts) and gallstone specimens were analyzed using FTIR Nicolet iS-5 Spectrometer from Thermo Fisher Scientific, USA. Thermo Scientific™ QCheck™ algorithm, embedded within the OMNIC™ software, was used to identify the unique spectral fingerprint of the patient samples to match with known, standard material. Matching of > 75% was considered acceptable. Validation for accuracy of the library was performed for twenty analyzed gallstones at an international reference lab. RESULTS: Concerted search analysis was performed against the developed GSL consisting of 71 "pure component" spectrum divided into 5 types to generate the library. For the Gallstone Real Patient Library (GRPL), 117 patient samples were analyzed. Ninety-eight gall stones (83.8%) out of 117 stones matched with the developed GSL. Majority stones were mixed stones (95.92%), with cholesterol being the primary component (91.83%). Results of the developed library were 100% in agreement with the reports received from the external reference lab. CONCLUSIONS: The library developed displayed good consistency and can be used for detection of gallstone composition in Pakistan and replace the traditional labor- and time-intensive chemical method of gallstone analysis.

Perspective on newborn screening (NBS): Evidence sharing on conditions to be included in NBS in Pakistan.

Newborn screening aims at detecting treatable disorders early so that the treatment can be initiated to prevent mortality and morbidity. Such programmes are well established in most developed countries, and all newborns are screened for selected metabolic, endocrine and other disorders based on disease epidemiology, testing and treatment availability, efficiency and cost-effectiveness. Even in developing countries, such screening programmes are initiated using heel prick capillary blood collected on filter paper. The current narrative review was planned to provide a perspective with evidence in favour of starting newborn screening for different disorders. The programme project should be initiated nationwide, taking one disorder, congenital hypothyroidism, as the prototype and a newborn screening panel can then be extended to include other disorders. A task force should be set up to recommend disorders to be included in the panel, develop the national plan policies, and define procedures to strengthen the testing.

Effect of physical activity and vitamin D compared with vitamin D alone on muscle strength, back flexibility and aerobic activity in patients with chronic kidney disease: A comparative study from Pakistan.

BACKGROUND AND OBJECTIVES: To compare the differences in musculoskeletal health with vitamin D alone in comparison with vitamin D with physical activity (PA) among chronic kidney disease (CKD) patients. METHODS AND STUDY DESIGN: An open labeled, randomized, controlled trial was conducted at two tertiary care centers in Pakistan. Patients with CKD stage 2-4 and vitamin D deficiency (<20 ng/mL) were recruited in the trial. Both the arms were given oral vitamin D (cholecalciferol) drops (4000 IU) once daily for three months. One arm received only vitamin D (VD arm), while the second arm received vitamin D along with PA (VDPA arm). RESULTS: Of the 1,235 CKD stage 2-4 subjects contacted, forty-six subjects were enrolled. Eighteen were assigned to VD arm and twenty-eight were assigned to VDPA arm. Between groups comparison shows that bicep strength increases from 15 to 17 kg. Likewise, back flexibility and aerobic fitness also increased among those who receive vitamin D and physical activity, however these differences were not statistically significant (p>0.05). Sensitivity analysis within group comparison shows rise of bicep strength from 13.8 kg to 15.2 kg in the VD alone arm (p=0.05); however, in the VDPA arm, there is a greater difference of 14.3 kg to 17.2 kg (p<0.001). CONCLUSIONS: Targeted PA among CKD patients has potential to improve bicep strength and back flexibility. However, as the sample size was small, further studies would be required to suggest whether a PA should be included as part of the treatment regimen.

Fostering teaching-learning through workplace based assessment in postgraduate chemical pathology residency program using virtual learning environment.

BACKGROUND: The principle of workplace based assessment (WBA) is to assess trainees at work with feedback integrated into the program simultaneously. A student driven WBA model was introduced and perception evaluation of this teaching method was done subsequently by taking feedback from the faculty as well as the postgraduate trainees (PGs) of a residency program. METHODS: Descriptive multimethod study was conducted. A WBA program was designed for PGs in Chemical Pathology on Moodle and forms utilized were case-based discussion (CBD), direct observation of practical skills (DOPS) and evaluation of clinical events (ECE). Consented assessors and PGs were trained on WBA through a workshop. Pretest and posttest to assess PGs knowledge before and after WBA were conducted. Every time a WBA form was filled, perception of PGs and assessors towards WBA, time taken to conduct single WBA and feedback were recorded. Faculty and PGs qualitative feedback on perception of WBA was taken via interviews. WBA tools data and qualitative feedback were used to evaluate the acceptability and feasibility of the new tools. RESULTS: Six eligible PGs and seventeen assessors participated in this study. A total of 79 CBDs (assessors n = 7 and PGs n = 6), 12 ECEs (assessors n = 6 and PGs n = 5), and 20 DOPS (assessors n = 6 and PGs n = 6) were documented. PGs average pretest score was 55.6%, which was improved to 96.4% in posttest; p value< 0.05. Scores of annual assessment before and after implementation of WBA also showed significant improvement, p value 0.039, Overall mean time taken to evaluate PG's was 12.6 ± 9.9 min and feedback time 9.2 ± 7.4 min. Mean WBA process satisfaction of assessors and PGs on Likert scale of 1 to 10 was 8 ± 1 and 8.3 ± 0.8 respectively. CONCLUSION: Both assessors and fellows were satisfied with introduction and implementation of WBA. It gave the fellows opportunity to interact with assessors more often and learn from their rich experience. Gain in knowledge of PGs was identified from the statistically significant improvement in PGs' assessment scores after WBA implementation.

Retrospective Study of Patients with Hyperphenylalaninemia- Experience from a Tertiary Care Center in Pakistan.

OBJECTIVE: To assess the clinical and biochemical features as well as outcome of hyperphenylalaninemia patients. Methods: The descriptive retrospective study was conducted at the Aga Khan University Hospital, Karachi, and comprised data from January 2013 to February 2017 of plasma amino acid analysed at the Biochemical Genetic Laboratory of patients with phenylalanine levels >120 umol/L. Medical charts of patients registered with the Metabolic Clinics were reviewed, while outside referrals were contacted by telephone to collect data on a pre-structured questionnaire. Data was analysed using SPSS 21. RESULTS: Of the 18 patients, 13(72%) were males. Overall median age was 606 days (interquartile range: 761) and median phenylalanine levels were 1280 (interquartile range: 935) umol/L. Phenylalanine hydroxylase deficiency was present in 5(28%) patients while 3(16.6%) had defects in the metabolism or regeneration of tetrahydrobiopterin. The most common clinical features was intellectual deficit and seizures 14(78%) each, followed by lighter hair colour 10(55.5%) and hypotonia 11(61%). High treatment cost was the leading reason for cessation of therapy in 7(39%) followed by refusal by patient's family 5(28%). CONCLUSIONS: Most hyperphenylalaninemia cases were diagnosed late when intellectual disability had already developed.

Inherited metabolic disorders presenting as hypoxic ischaemic encephalopathy: A case series of patients presenting at a tertiary care hospital in Pakistan.

In spite of the efforts and interventions by the Government of Pakistan and The World Health Organization, the neonatal mortality in Pakistan has declined by only 0.9% as compared to the global average decline of 2.1% between 2000 and 2010. This has resulted in failure to achieve the global Millennium Development Goal 4. Hypoxic-ischaemic encephalopathy, still birth, sepsis, pneumonia, diarrhoea and birth defects are commonly attributed as leading causes of neonatal mortality in Pakistan. Inherited metabolic disorders often present at the time of birth or the first few days of life. The clinical presentation of the inherited metabolic disorders including hypotonia, seizure and lactic acidosis overlap with clinical features of hypoxic-ischaemic encephalopathy and sepsis. Thus, these disorders are often either missed or wrongly diagnosed as hypoxicischaemic encephalopathy or sepsis unless the physicians actively investigate for the underlying inherited metabolic disorders. We present 4 neonates who had received the diagnosis of hypoxic-ischaemic encephalopathy and eventually were diagnosed to have various inherited metabolic disorders. Neonates with sepsis and hypoxic-ischaemic encephalopathy-like clinical presentation should be evaluated for inherited metabolic disorders.

Association between environmental tobacco smoke and dental caries amongst 5-14 years old children in Karachi, Pakistan.

OBJECTIVE: To determine the association between environmental tobacco smoke and dental caries. METHODS: This cross-sectional study was conducted in peri-urban and urban areas of Karachi, from February to August 2014, and comprised children aged 5-14 years. A pre-coded questionnaire for environmental tobacco smoke and food frequency questionnaire for dietary habits were used. Dental examination of children was done to detect caries. Cox-proportional hazard algorithm was used to measure the association of environmental tobacco smoke with dental caries at multivariable level. STATA version 12.0 was used for statistical analysis. RESULTS: Of the 500 children, 250(50%) each were from peri-urban and urban localities. The prevalence of dental caries was 336(67.2%).Family members of 154(30.8%) participants reported smoking. After adjusting for junk food intake, in-between meals, age, plaque index, dental visits and socio-economic status, the association between environmental tobacco smoke and dental caries remained statistically significant (p<0.05). Compared to non-exposed children, the adjusted prevalence ratio was 1.25 (95% confidence interval: 1.08-1.46) and 1.36 (95% confidence interval: 1.09-1.70) for children with < 30 minutes and >30 minutes of environmental tobacco smoke exposure, respectively. CONCLUSIONS: Environmental tobacco smoke was found to be associated with dental caries.

Diagnostic dilemma of patients with methylmalonic aciduria: Experience from a tertiary care centre in Pakistan.

OBJECTIVE: To determine the frequency of disorders leading to methylmalonic acidurias. METHODS: This cross-sectional study was conducted from January 2013 to April 2016 at the Aga Khan University Hospital, Karachi, and comprised patients diagnosed with methylmalonic acidurias based on urine organic acid analysis. Clinical history and biochemical data was collected from the biochemical genetics laboratory requisition forms. Organic acid chromatograms of all the subjects were critically reviewed by a biochemical pathologist and a metabolic physician. For assessing the clinical outcome, medical charts of the patients were reviewed. SPSS 19 was used for data analysis. RESULTS: Of the 1,778 patients 50(2.81%) were detected with methylmalonic acidurias. After excluding patients with non-significant peaks of methylmalonic acidemia, 41(2.31%) were included in the final analysis. Of these, 20(48.7%) were females, while the overall median age was 11.5 months (interquartile range: 6-41.5). On stratification by type of disorders leading to methylmalonic acidurias, 9(22%) had methylmalonic acidemia, 12(29%) had Cobalamin-related remethylation disorders, nonspecific methylmalonic acidurias in 16(39%), while 2(5%) each had succinyl coenzyme A synthetase and Vitamin B12 deficiency. respectively. CONCLUSIONS: Screening tests, including urine organic acid, provided valuable clues to the aetiology of methylmalonic acidurias.

Burden of Zinc (Zn) deficiency: A high volume clinical laboratory data analysis.

A cross-sectional survey was done at the Section of Chemical Pathology, Department of Pathology and Laboratory Medicine, AKUH, Karachi, to determine frequency of Zn deficiency, the second major micronutrient deficiency in developing countries, in subjects coming for investigations. Data of plasma Zn, from Jan 2013 to Dec 2014 was analyzed and deficiency was labeled at <65ug/dl. Total 469 subjects were tested for plasma Zn levels over the period of 24 months and complete information was available for 422 subjects, included in final analysis. Median age of subjects was 13 yrs (IQR 7-35) and 57% were male. Mean plasma Zn levels of the total subjects were 105±42.6ug/dl. Out of the total subjects 13.5% (n=57) had Zn deficiency (mean plasma Zn 52±11 ug/dl), and most of them were of age group 6-16yrs. These findings advocate that diet of Pakistani children should be revised in order to provide sufficient amounts of Zn.

IDENTIFYING PARATHYROID HORMONE DISORDERS AND THEIR PHENOTYPES THROUGH A BONE HEALTH SCREENING PANEL: IT'S NOT SIMPLE VITAMIN D DEFICIENCY!

OBJECTIVES: To determine the utility of bone health screening panels in identifying disorders of parathyroid gland secretions. METHODS: A retrospective analysis of biochemical parameters in a bone health screening panel (BHSP) was conducted. Low and high cutoffs were applied to determine hypofunctioning and hyperfunctioning conditions related to parathyroid hormone. Clinical phenotypes of parathyroid gland abnormalities were determined using a combination of levels of calcium, 25-hydroxyvitamin D, and intact parathyroid hormone (iPTH). A PTH nomogram was applied to calculate the maximum expected PTH for existing levels of 25-hydroxyvitamin D. Medical records of patients were reviewed for clinical validation of biochemical findings. RESULTS: Sixty-eight percent of subjects showed abnormal PTH secretion. Primary hyper- and hypoparathyroidism were detected in 1% (n = 5) and 0.4% (n = 2) of subjects, respectively. Normocalcemic hyperparathyroidism and hypercalcemia with inappropriately high-normal PTH were identified in 8.5% (n = 37) and 2% (n = 10) of subjects, respectively. All subjects with primary and normocalcemic hyperparathyroidism had higher measured PTH than calculated maximum PTH using the PTH nomogram. Secondary hyperparathyroidism and functional hypoparathyroidism were present in 18% (n = 88) and 39% (n = 194) of subjects, respectively. High prevalence of bone pains, renal stones, and low bone mineral density were identified in patients with abnormal PTH secretion. CONCLUSION: Panel testing is useful in early diagnosis of metabolic bone disorders related to PTH. A BHSP helps identify normocalcemic hyperparathyroidism and hypercalcemia with inappropriately high PTH. ABBREVIATIONS: 25OHD = 25-hydroxyvitamin D AKUH = Aga Khan University Hospital BHSP = bone health screening panel iPTH = intact parathyroid hormone maxPTH = maximum parathyroid hormone MBD = metabolic bone disease NCHPT = normocalcemic hyperparathyroidism PHPT = primary hyperparathyroidism PTH = parathyroid hormone SHPT = secondary hyperparathyroidism VDD = vitamin D deficiency.

Error identification in a high-volume clinical chemistry laboratory: Five-year experience.

INTRODUCTION: Quality indicators for assessing the performance of a laboratory require a systematic and continuous approach in collecting and analyzing data. The aim of this study was to determine the frequency of errors utilizing the quality indicators in a clinical chemistry laboratory and to convert errors to the Sigma scale. MATERIALS AND METHODS: Five-year quality indicator data of a clinical chemistry laboratory was evaluated to describe the frequency of errors. An 'error' was defined as a defect during the entire testing process from the time requisition was raised and phlebotomy was done until the result dispatch. An indicator with a Sigma value of 4 was considered good but a process for which the Sigma value was 5 (i.e. 99.977% error-free) was considered well controlled. RESULTS: In the five-year period, a total of 6,792,020 specimens were received in the laboratory. Among a total of 17,631,834 analyses, 15.5% were from within hospital. Total error rate was 0.45% and of all the quality indicators used in this study the average Sigma level was 5.2. Three indicators - visible hemolysis, failure of proficiency testing and delay in stat tests - were below 5 on the Sigma scale and highlight the need to rigorously monitor these processes. CONCLUSIONS: Using Six Sigma metrics quality in a clinical laboratory can be monitored more effectively and it can set benchmarks for improving efficiency.

An audit of clinical laboratory data of 25 [OH]D at Aga Khan University as reflecting vitamin D deficiency in Pakistan.

OBJECTIVE: To estimate the burden of Vitamin D deficiency in people from different geographical areas of Pakistan. METHODS: The retrospective study was conducted at the Aga Khan University Hospital, Karachi, and comprised specimens of subjects tested for 25-hydroxy D at the clinical laboratory between September 2010 and September 2011. Serum samples received from the phlebotomy centres all over Pakistan and the main laboratory in Karachi were included. SPSS 19 was used for statistical analysis. RESULTS: Of the 60937 specimens in the study, 18721(30.7%) related to men. The overall mean age was 40.5±19.7 years, and median 25-hydroxy D level was 13.5ng/ml (interquartile range: 25.1-7.4 ng/ml).Overall, mean log 25-hydroxy D was 1.14±0.39ng/ml (median: 13.5ng/ml; interquartile range: 25.1-7.4 ng/ml). Out of the total, 40279(66.1%) subjects were vitamin D-deficient. CONCLUSIONS: Vitamin D deficiency was common among the subjects.

Ionized calcium measurement in serum and plasma by ion selective electrodes: comparison of measured and calculated parameters.

To determine the concentration of ionized calcium (iCa) collected in lithium heparin and gel tubes and to correlate the measured iCa with calculated iCa. Anaerobic fasting blood samples were simultaneously collected from healthy laboratory workers in lithium-heparin and gel tubes. iCa, pH, total calcium (CaT), total protein and albumin were measured. Ionized Ca was calculated with albumin and globulin values using an appropriate formula. Mean iCa in gel tubes showed a positive constant bias of 0.08 mmol/L (p < 0.001) when compared to lithium heparin results. The mean pH of blood taken in lithium heparin and gel tubes was non-significant (p = 0.3). Measured iCa poorly correlated with CaT (r = 0.2, p = 0.1) and calculated iCa (r = 0.2, p = 0.2). To evaluate the calcium status direct measurement of iCa must be done rather than using the formulae for iCa determination. In addition serum is recommended as the choice of sample for iCa determination in comparison to plasma samples.