Acid freezes uricolysis in addition to glycolysis: Utility of fluoride-EDTA-citrate tube in the measurement of uric acid for patients administered rasburicase

Background In samples from patients administered rasburicase, ex vivo uricolysis leads to spuriously low uric acid results. The manufacturer’s recommendation of storing the sample in ice-water until analysis, however, does not fully arrest uricolysis. Since uricase activity is affected by pH and metal chelators, we assessed uricolysis inhibition in sodium fluoride-ethylenediaminetetraacetic acid (EDTA)-citrate sample tube (FC Mix tube, Greiner) used primarily for plasma glucose. Method A serum pool was spiked with rasburicase and uric acid measured at 15, 45, 90, 150, 240 and 1080 min in a lithium heparin tube in ice-water, plain tube at room temperature (RT), EDTA tube at RT, FC Mix tube in ice-water, FC Mix tube at RT and FC Mix tube at RT prepared by dissolving FC Mix in serum. Results The rate of urate decay was lowest in the FC Mix tube independent of temperature, then lithium heparin tube in ice-water, then EDTA tube at RT and highest in the plain tube at RT. Uric acid concentrations in the prepared FC Mix tube at RT and heparin tube in ice-water were, respectively, 98.2% and 93.8% of control values at 90 min, 97.1% and 89.3% of control values at 4 h, and remained higher in the prepared FC Mix tube at all time points. Conclusion NaF-EDTA-citrate mixture largely arrested rasburicase mediated ex vivo uricolysis without the need for sample cooling. We propose that sample tubes containing NaF-EDTA-citrate be used for the measurement of uric acid in patients administered rasburicase.

THE EFFECTS OF EXERCISE MODELS ON HEMATOLOGICAL PROFILE AND MEET PHYSICAL QUALITY IN GARUT SHEEP

The objective of this study was to evaluate the effect of exercise levels on the hematological and metabolical status, and carcass quality of Garut sheep. In total 24 of Garut sheep used in this study. The levels of exercise contain three treatments namely non exercise, semi exercise and exercise. Blood collection through the jugular vein of sheep samples was carried out at the end of the study, using a 5 mL EDTA tube to measure the haematological condition of the animal sample. Likewise, collecting muscle samples to determine the physical quality of meat.The data was analyzed by analysis of variance and continued by Duncan test for a post hoc test. The results showed that the models of exercise were not significantly (P0.05) affected to hematological level and also carccas quality including juiceness, tenderness, drip loss and water holding capacity. Organoleptic sensory using semiexercise was more favourable meat compare to exercise and non exercise. While for the colour of meat, non exercise treatment was significantly (P0.05) affected to the colour of meat compare to exercise and semi exercise. Furthermore the semi exercise was the best treatment to produce fresh taste meat with good sensory characteristic.

Human Plasmodium in Domestic Animals in West Sumba and Fakfak, Indonesia

Background: Although maximum efforts have been made, malaria in several areas in Indonesia is still high. This study aims to detect the possibility of a Plasmodium reservoir in domestic animals in endemic malaria areas.Methods: Blood from the domestic animal was collected by EDTA tube, smeared and stained by Giemsa for detecting Plasmodium microscopically. Ten µl blood from EDTA tube dripped into filter paper for Plasmodium DNA capture. Nested PCR was used for the molecular detection of parasites, and DNA was sequenced from PCR products to ascertain Plasmodium species.Result: A total of 208 and 62 animal blood samples were collected from Gaura and Fakfak villages. Thirty-two of 270 animals contained P. falciparum or P. vivax, and all are from Gaura village. The percentage of Plasmodium in buffalo, horse, goat, and dog is 20.7%, 14.3%, 5.8%, 16.7%, respectively. Neither P. knowlesi found in all samples, nor parasite detected in 18 pig blood samples.Conclusion: Human Plasmodium exists in domestic animals in Indonesia. This finding may partly explain the persistence of the high prevalence of malaria in some endemic areas in Indonesia and may affect public health and malaria control strategy.

Reference Intervals for and the Effects of Sample Handling and Sex on Rotational Thromboelastometry in Healthy Adult Pigs

Accurate assessment of coagulation in porcine studies is essential. We sought to establish normal values for porcine rotational thromboelastometry (ROTEM) according to the American Society for Veterinary Clinical Pathology guidelines and to assess the effects of various preanalytical parameters on those measurements. Healthy Yorkshire-cross pigs (n = 81; 46 males and 35 females) were anesthetized. By using a 18-gauge needle attached to a vacuum phlebotomy tube, blood was acquired from the cranial vena cava. Tubes were filled in the following order: evacuation clot tube, EDTA tube, heparin tube, and 2 citrate tubes. The citrate tubes were randomly assigned to 30 min with or without constant agitation on a rocker. The following parameters were reported according to the manufacturer's recommendations: clotting time, clot formation time, α, (tangent to the clot formation curve when the clot firmness is 20 mm), clot firmness after 10 and 20 min, maximal clot firmness, maximum lysis, and lysis indexes at 30 and 45 min. Reference intervals were reported as mean ± 2 SD (parametric distribution) or 2.5th and 97.5th percentile of the population's results (nonparametric distribution). The effects of sex, sampling order, and agitation on ROTEM results were analyzed through linear regression. Neither sex nor sample agitation influenced any of the ROTEM parameters. Combined reference intervals were established for each ROTEM parameter by pooling data from the nonagitated tubes for both male and female pigs. This study is the first to establish ROTEM reference intervals from a large number of male and female adult Yorkshire-cross pigs and to provide a detailed description of preanalytical sample processing.

Profil Darah Rusa Totol (Axis axis) Betina Sehat di Pusat Inovasi Agroteknologi (PIAT), Universitas Gadjah Mada

Center for Agrotechnology Innovation, Gadjah Mada University (PIAT), one of the place for development and breeding ground of spotted deer in Yogyakarta, Indonesia. The study of blood profiles was intended to allow veterinarian to understand the normal profile of the spotted deer. Fifteen healthy spotted deer belonging to PIAT were used as experimental animals. The deer were drawn its blood through the jugular vein without the use of anaesthesia. Blood was then accommodated in an EDTA tube, centrifuged at 2500 RPM and analyzed using a Mindray BC-2800 haematology analyzer machine. Based on the results of the study of the blood it were known that Haemoglobine (Hb) was: 11.5 ± 1.703 g/dl, Red blood cells (RBC) was: 9.3 ± 3.580 106/ml, Packed cell volume (PCV) was: 30.8 ± 6.035 %, Mean corpuscular Volume (MCV) was: 36.8 ± 11.102 fl, Mean corpuscular haemoglobine (MCH) was: 15.0 ± 7.313 pg, Mean corpuscular haemoglobine concentration (MCHC) was: 40.0 ± 14.657 g/dl, White blood cells (WBC) was: 6.4 ± 3.096 103/ml , Neutrophils was: 43.4 ± 21.646, % Basophils was: 0.2 ± 0.168 %, eosinophils was: 0.4 ± 0.447 %, lymphocytes was: 53.4 ± 21.546 % and monocytes was: 2.6 ± 2.394 %. From all of the datas it indicated that blood profiles of PIAT’s spotted deers are different from similar spotted deer blood profiles in India.

The Diagnostic Value of the Glucose Curve in Individuals with and without Diabetes

Aim: Of this study is to demonstrate the importance of glucose curve test in monitoring pre and post-meal variation in diabetic and normal individuals. Methodology: The individuals subjected to this study mainly grouped in two categories the (DM2 group) and the (Control group), they instructed to came fasting at which blood sample will be collected in EDTA and blank tube then after 30 min. the first post-prandial blood sample collected and then after every 1,2,3,4,5,6,7,8 hours blood sample collected subsequently, then serum separated from each sample (except the EDTA tube) analysed biochemically for glucose and glycated haemoglobin HbA1c (from EDTA tube). Result: We found that, the calculated glucose based on mean glycated haemoglobin HbA1c% results underestimate the real concentrations all over the glucose curve in control group but in DM2 group it underestimate the mean and some actually measured concentration in some points of the curve which adds more burden on the diabetic patient and the responsibility of adjusting the dose and time of administration. Conclusion: from our prospect we recommend the use of blood glucose curve as a monitoring and diagnostic tool generally for glucose metabolism in normal, pre-diabetic, diabetic and uncontrolled diabetic patients before and during therapeutic conditions.

Assessment of dried blood spots for DNA methylation profiling

Background: DNA methylation reflects health-related environmental exposures and genetic risk, providing insights into aetiological mechanisms and potentially predicting disease onset, progression and treatment response. An increasingly recognised need for large-scale, longitudinally-profiled samples collected world-wide has made the development of efficient and straightforward sample collection and storage procedures a pressing issue. An alternative to the low-temperature storage of EDTA tubes of venous blood samples, which are frequently the source of the DNA used in such studies, is to collect and store at room temperature blood samples using purpose built filter paper, such as Whatman FTA® cards. Our goal was to determine whether DNA stored in this manner can be used to generate DNA methylation profiles comparable to those generated using blood samples frozen in EDTA tubes. Methods: DNA methylation profiles were obtained from matched EDTA tube and Whatman FTA® card whole-blood samples from 62 Generation Scotland: Scottish Family Health Study participants using the Infinium HumanMethylation450 BeadChip. Multiple quality control procedures were implemented, the relationship between the two sample types assessed, and epigenome-wide association studies (EWASs) performed for smoking status, age and the interaction between these variables and sample storage method. Results: Dried blood spot (DBS) DNA methylation profiles were of good quality and DNA methylation profiles from matched DBS and EDTA tube samples were highly correlated (mean r = 0.991) and could distinguish between participants. EWASs replicated established associations for smoking and age, with no evidence for moderation by storage method. Conclusions: Our results support the use of Whatman FTA® cards for collecting and storing blood samples for DNA methylation profiling. This approach is likely to be particularly beneficial for large-scale studies and those carried out in areas where freezer access is limited. Furthermore, our results will inform consideration of the use of newborn heel prick DBSs for research use.

Assessment of Dried Blood Spots for DNA Methylation Profiling

BackgroundDNA methylation reflect health-related environmental exposures and genetic risk, providing insights into aetiological mechanisms and potentially predicting disease onset, progression and treatment response. An increasingly recognised need for large-scale, longitudinally-profiled samples collected world-wide has made the development of efficient and straightforward sample collection and storage procedures a pressing issue. An alternative to the low-temperature storage of EDTA tubes of venous blood samples, which are frequently the source of the DNA used in such studies, is to collect and store at room temperature blood samples using filter paper engineered for the purpose, such as Whatman FTA®cards. Our goal was to determine whether DNA stored in this manner can be used to generate DNA methylation profiles comparable to those generated using blood samples frozen in EDTA tubes.MethodsDNA methylation profiles were obtained from matched EDTA tube and Whatman FTA®card whole-blood samples from 62 Generation Scotland: Scottish Family Health Study participants using the Infinium HumanMethylation450 BeadChip. Multiple quality control procedures were implemented, the relationship between the two sample types assessed, and EWASs performed for smoking status, age and the interaction between these variables and sample storage method. Results: Dried blood spot (DBS) DNA methylation profiles were of good quality and DNA methylation profiles from matched DBS and EDTA tube samples were highly correlated (mean r = 0.991) and could distinguish between participants. EWASs replicated established associations for smoking and age, with no evidence for moderation by storage method.ConclusionsOur results support the use of Whatman FTA®cards for collecting and storing blood samples for DNA methylation profiling. This approach is likely to be particularly beneficial for large-scale studies and those carried out in areas where freezer access is limited. Furthermore, our results will inform consideration of the use of newborn heel prick DBSs for research use.

KESAHIHAN DIAGNOSTIK HEMOGLOBIN RETIKULOSIT UNTUK DETEKSI DEFISIENSI ZAT BESI DI KEHAMILAN (Diagnostic Validity of Reticulocyte Hemoglobin for Iron Deficiency Detection in Pregnancy)

Entering the second trimester of pregnancy, more iron is required due to the increase in erythrocyte mass, plasma volume andthe development of fetus as well as chorion. Iron is needed the most in the third trimester. The existing hematological iron stageparameters can only detect iron deficiency in the latest stage. The aim of this study was to know the assessment validity of Ret-Heexamination as a new parameter to diagnose iron deficiency in pregnant women with anemia, as well as a screening tool for those interm pregnancy without anemia. The research design was cross sectional. The subjects were women in term pregnancy, gathered fromPKU Muhammadiyah Hospital, Bantul Yogyakarta from May to November 2013. A seven (7) mL blood sample was taken from thecubital vein of the subjects. Two mL of the sample was tested for routine hematological examination using an EDTA tube, while theRet-He was assessed using an automatic hematological instrument Sysmex XT-2000-i (Symex Corporation, Kobe, Japan). The serumof the remaining five (5) mL was used to check the serum iron and TIBC to obtain the saturation value (Tsat) using Cobas analyzerC501 (Roche Diagnostics, Germany), while the serum ferritin (SF) was examined using Minividas. The subjects were classified into two(2) groups based on the Hb levels, namely: anemia (Hb<11 g/dL) and those who did not (Hb≥11 g/dL). Furthermore, they were alsoclassified into two (2) groups based on transferrin saturation values: iron deficient (Tsat <9%) and normal (Tsat ≥9%). From 291subjects, 59 (20.3%) were found to have anemia and 232 (79.7%) did not. The cut off value of Ret-He to diagnose iron deficiency inpregnant women with anemia was 29.8 pg (82% sensitivity and 72% specificity). Meanwhile, the cut-off value of Ret-He for irondeficiency screening in pregnant women without anemia was 29.8 pg, with a sensitivity and specificity of 92% and 87% respectively.The Ret-He holds a good diagnostic validity to detect iron deficiency in pregnancy, with or without anemia.