10-mL Tubes Research Articles

Statistical comparisons of 0.08 g/100 ml fortified blood alcohol samples from 6-ml and 10-ml gray-top tubes.

This experiment supplements the study, "Statistical comparisons of blood alcohol samples from 6-ml and 10-ml grey-top tubes". The initial study analyzed fortified samples for blood alcohol concentration (BAC) using two sizes of gray-top tubes: a 10-ml tube containing a nominal 1% sodium fluoride (NaF), a preservative, and a 6-ml tube containing 0.25% NaF, using the variables of time, storage temperature, fill volume, and concentration. From that study, paired t-tests determined no difference in BAC from data sets of the two tube sizes, however, Analysis of Variance, or ANOVA, testing determined the two tube sizes yielded different results at 0.08 g/100 ml. To investigate this potential disparity, this study fortified new samples at 0.08 g/100 and focused on two variables: fill volume and storage duration, while increasing the sample population (to n=8). For this study, the two tube types yielded equivalent concentrations under the majority of conditions studied. Differences between the two tube types were found using paired t-test for the high-volume samples on Days 7 and 30; ANOVA yielded the same results but also determined one additional statistical difference for the Day 30 low-volume samples. However, the differences observed between tube sizes fall within standard deviation ranges established for the analytical method precision profile, indicating statistical differences are not analytically significant. Further studies are needed to investigate the comparability of the tubes under real-world conditions, under which oxygen and/or additives are not added during sample preparation steps or by usage of blood bank products.

Simultaneous determination of monochloropropanediol esters and glycidyl esters in vegetable oils by acidic transesterification-gas chromatography-mass spectrometry

建立了一种气相色谱-质谱同时测定植物油中3-氯丙二醇酯、2-氯丙二醇酯和缩水甘油酯的方法。称取0.25 g样品,依次加入内标工作液、四氢呋喃和酸性溴化钠溶液,50 ℃水浴反应15 min,加入6 g/L碳酸氢钠溶液终止反应,使用正己烷提取,上层液经氮气吹干后用四氢呋喃溶解。随后加入1.8%(v/v)硫酸-甲醇溶液于40 ℃恒温水浴中反应16 h,加入饱和碳酸氢钠溶液终止反应。样液再经过净化、衍生、提取、氮吹后,以1 mL正己烷定容,过膜,进样测定。采用毛细管气相色谱柱DB-5MS(30 m×0.25 mm×1 μm)分离,程序升温,电子轰击电离(EI)源检测,在选择离子扫描模式下,以保留时间和特征离子信息进行定性分析,内标法定量。结果表明,3-氯丙二醇酯、2-氯丙二醇酯和缩水甘油酯在0.01~0.80 mg/L范围内线性关系良好,相关系数(r2)均在0.999以上,方法的检出限(S/N=3)和定量限(S/N=10)分别为25、25、20 μg/kg和75、75、60 μg/kg。选取4种不同基质类型的样品,在低、中、高3个不同添加水平下的平均回收率为89.0%~98.7%,相对标准偏差(RSD)在2.05%~7.81%之间。采用该方法测定了市售112份植物油样本,其中有84份样本检出3-氯丙二醇酯、2-氯丙二醇酯和缩水甘油酯。与已建立的国家标准方法(GB 5009.191-2016)和行业标准方法(SN/T 5220-2019)相比,该方法所采用的酸性酯交换法可避免副反应(碱性条件下3-氯丙二醇、2-氯丙二醇及3-溴丙二醇向游离态缩水甘油转化)的发生,同时该方法也弥补了国家标准和行业标准无法对3-氯丙二醇酯、2-氯丙二醇酯和缩水甘油酯同时进行测定的缺失。该方法实验操作更高效,结果更准确、重复性更好,对我国植物油中3-氯丙二醇酯、2-氯丙二醇酯和缩水甘油酯污染水平的控制、检测标准的制定和生产工艺的优化具有一定的理论和现实意义。

Isobutylene contamination of blood collected in 10-ml evacuated blood collection tubes with gray conventional rubber stoppers.

Dual-column headspace gas chromatographic analysis with two flame-ionization detectors is a commonly used analytical technique for forensic blood ethanol quantitation. This technique is also applicable to the identification and quantitation of other volatile organic compounds such as methanol in biological samples. Compound identification by retention time is limited to those compounds with known retention times programmed into the instrument method. Historically, an early-eluting peak from an unidentified compound has been observed in both chromatograms from antemortem blood samples analyzed for ethanol concentration with this technique. The unidentified compound's retention time matches that of methanol on one column but not on the second column. This previously unidentified compound has been identified as isobutylene. The proposed source of the isobutylene contamination historically observed in antemortem blood samples collected in 10-ml gray-top blood collection tubes is the conventional rubber stopper. Isobutylene was detected in deionized water stored in each of the seven lots of 10-ml blood tubes tested; the expiration dates of the tubes tested spanned the years 2002-2022. Misidentification of isobutylene as methanol is possible when using a single-column gas chromatographic system. The presence of isobutylene in blood collected in a gray-top collection tube does not represent laboratory contamination, is not an interferent with blood ethanol quantitation, and does not affect the ethanol concentration in the blood. A 0.150g/dl aqueous ethanol standard was stored in a gray-top tube to evaluate the potential impact of isobutylene on ethanol quantitation. The solution's average ethanol concentration measured after storage was 0.150g/dl.

β-Amyloid in Cerebrospinal Fluid: How to Keep It Floating (Not Sticking) by Standardization of Preanalytic Processes and Collection Tubes.

Phosphorylated tau (pTau), total tau (tTau), and β-amyloid (Aβ) are established cerebrospinal fluid (CSF) biomarkers used to help diagnose Alzheimer disease. Preanalytic workups of CSF samples lack harmonization, making interlaboratory comparison of these biomarkers challenging. The Aβ adsorbs to sample tubes, yielding underestimated concentrations, and may result in false Alzheimerdisease diagnosis. Our primary aim was to compare Aβ recovery across multiple polypropylene tubes and to test the stability of tTau, pTau, and Aβ in the best performing tube. Eight polypropylene tubes were tested using 3 CSF pools with Aβ concentrations <500, 500-1000, and >1000 ng/L. All samples were analyzed in duplicate. Tubes were cut open to assess their different infrared adsorption spectra. Freshly drawn CSF from 14 patients was distributed into 4 Sarstedt 5-mL (no. 63.504.027; Sar5CSF) tubes, left at room temperature for up to 7 days, and analyzed for pTau, tTau, and Aβ by ELISA. Two Sarstedt 5-mL tubes and a Sarstedt 10-mL (Sar10CSF) tube showed significantly higher Aβ recovery at all 3 concentrations compared with the 5 other tubes. The infrared adsorption spectra of Sar10CSF and Sar5CSF tubes were practically identical, unlike the other tubes. No significant loss of pTau, tTau, and Aβ was observed in CSF left at room temperature for up to 7 days (P > 0.05). Recovery of Aβ from Sar5CSF tubes is equivalent to Aβ recovery from Sar10CSF tubes. Levels of pTau, tTau, and Aβ were stable for at least 7 days at room temperature but not at 37 °C.

Statistical comparisons of blood alcohol samples from 6-mL and 10-mL grey-top tubes.

Historically, blood alcohol concentration (BAC) studies utilized a 1% concentration of the preservative sodium fluoride (NaF), leaving an information gap supporting usage of lower concentrations of NaF to preserve ethanol. As many forensic laboratories utilize Becton, Dickinson and Company 6-mL gray-top tubes (0.25% NaF), statistical comparisons were conducted to determine whether significant differences exist between BAC values obtained from 6-mL tubes versus 10-mL tubes (1% NaF). Whole blood was spiked at three concentrations, (0.04, 0.08, and 0.15g/100mL) and aliquoted into tubes at "low," "medium," and "high" fill volumes. Tubes were split into refrigerated or ambient storage and analyzed after 1, 3, 5, 7, and 30days, using headspace gas chromatography. Each 6-mL and 10-mL tube pair, prepared, stored, and analyzed under identical conditions, was compared by t-test (95% confidence level). For refrigerated tubes, 32 of 45-tube pairs did not reject the null hypothesis (that 6-mL and 10-mL tubes yield equivalent BACs), and 31 of 45 ambient stored tube pairs did not reject the null hypothesis. Analysis of variance (ANOVA) found no significant differences between 6-mL and 10-mL gray-top tubes for 0.04 and 0.15g/100mL concentrations over 30days; significant differences were observed for 0.08g/100mL concentration tubes, which warrants further study. Paired t-tests of grouped samples found no significant differences between 6-mL and 10-mL tubes at any concentration.

Beryllium characterization in soils by selective extraction

In the present work, an analytical method has been developed for the determination of the amount of man-made beryllium metal and/or beryllium compounds in soil in addition to the amount of native beryllium that is present. The method is based on the premise that if high-fired beryllium oxide can be extracted efficiently, then beryllium metal and acid-soluble beryllium salts would also be extracted efficiently.The proposed method involves the addition of 9 mL of an aqueous solution of 5% (wt/vol) ammonium bifluoride (ABF) to a 10-mL tube with 250 mg of dried, sieved soil. The tube is placed in a convection oven at 70 ᵒC and is mechanically rocked for 60 min to mix the sample while it is being heated. Beryllium oxide, beryllium metal and beryllium salts would include virtually all of the anthropogenic (man-made) beryllium compounds that might contaminate soil. Compounds that may be naturally present in soil, such as beryllium silicates and beryllium aluminosilicates, are not extracted efficiently by ABF solution under these conditions. As a second step after the initial extraction of the sample of soil with ABF solution, the soil is completely dissolved in a mixture of strong acids and that acid solution is then analyzed to determine the residual beryllium.The ratio of the extractable to residual beryllium in the same aliquot of native soil can be measured for a soil type in a local region and then used to calculate the amount of anthropogenic beryllium in soil. Therefore, the method may have application in the measurement of beryllium-contaminated soil at sites where beryllium oxide, beryllium metal and/or beryllium salts may have been used.

SA96 - ASSOCIATION STUDY POLYMORPHISMS OF THE NEUREGULIN 1 (NRG1) GENE WITH SCHIZOPHRENIA

Background The dysfunction of Neuregulin 1 (NRG1) is one of the plausible hypotheses for the pathogenesis of schizophrenia. The Neuregulin 1 (NRG1) is located on chromosome 8p, as suggested by multiple linkage studies. The aim of this study is to clarify the contribution of polymorphisms of the Neuregulin 1 (NRG1) with schizophrenia. Methods After informed consent was obtained, 100 schizophrenia patients and 100 control subjects were enrolled in this study. All subjects were administered the Diagnostic Interview for Genetic Studies (DIGS) (National Institute of Mental Health-Molecular Genetics Initiative, 1992; Nurnberger et al., 1994) by a research assistant with extensive training in this interview. Blood samples were collected in anonymously identified 10-ml Vacutainer tubes (Becton Dickinson). DNA was prepared by a modified SDS/Proteinase K procedure (Gusells et al., 1979). We genotyped polymorphism Neuregulin 1 (NRG1) with the PCR-RFLP methods. The PCR products were digested by restricted enzyme. Results We observed a significant association between the polymorphism neuregulin 1 (NRG1) and the schizophrenia (Chi-Square Test P= 0.0449). Discussion The NRG1 gene was originally identified as a susceptibility gene for schizophrenia by using a combination of a linkage and association approaches based on microsatellite markers and then using SNPs after microsatellite at risk haplotypes were identified. We found there is the frequency of the polymorphism of Neuregulin 1 (NRG1) was significantly increased in schizophrenia patients. This allelic association suggests that the functional polymorphism Neuregulin 1 (NRG1) may play a role in susceptibility to schizophrenia. Further study with larger sample sizes is required.

The effect of age, gender, and time between blood draw and start of centrifugation on the size outcomes of platelet-rich fibrin (PRF) membranes.

Platelet-rich fibrin (PRF) has been utilized in regenerative dentistry as a supra-physiological concentrate of autologous growth factors capable of stimulating tissue regeneration. Due to the variability in the macroscopic morphology/size of PRF membranes observed between patients, we were interested in studying the effects of patient age, gender, and time between blood draw and the start of centrifugation on the size outcomes of PRF membranes. Despite PRF therapy being increasingly more popular in private practice, to date, no study has investigated the effects of the delay between blood draw and the start of centrifugation in a clinical setting. A total of 60 patients enrolled in this study were divided into 6 groups of 10 patients each, including male and female patients categorized into age groups 21-40, 41-60, and 61-80years. From each patient, a total of five PRF membranes were fabricated from 10-mL tubes following centrifugation starting after 0, 30, 60, 90, and 120s. In total, 300 PRF membranes were produced in this study to investigate the effects of patient age, gender, and time on the size outcomes of PRF membranes. A longer delay by the clinician before starting centrifugation following blood draw led to a smaller final size of PRF membranes. At 90s following blood draw, a significant (13%) reduction in PRF membrane size was observed. After 120s, a significant (23%) reduction was observed. Additionally, female patients had on average 17% larger membranes compared to men (p < 0.05, 300 samples). Lastly, the size outcomes of the PRF membranes was largest in patients aged 61-80, followed by those aged 41-60 and 21-40. However, no statistically significant differences in PRF membrane sizes were reported between age groups. The time at which a centrifugation procedure begins following blood draw is critical to optimize the size outcomes of PRF membranes. In general, approximately 15s is required per tube to harvest 9-10cc of blood. Therefore, a 60- to 90-s interval between blood draw and the start of centrifugation should be a parameter that is respected by clinicians to avoid significant changes in the macroscopic morphology/size of fabricated PRF membranes. Furthermore, females and older patients produced larger membranes, likely due to lower red blood cell counts derived from their peripheral blood. The findings from the present study demonstrate that on average, a clinician has approximately 60-90s between blood draw and the start of the centrifugation cycle to produce standard-sized PRF membranes. Shortly thereafter, a significant reduction in size is observed. Additionally, females and older patients were found to produce larger PRF membranes. Centrifugation protocols may therefore be adapted accordingly.

T186. ASSOCIATION BETWEEN POLYMORPHISMS OF THE NEUREGULIN 1 (NRG1) GENE AND SCHIZOPHRENIA

BackgroundThe dysfunction of neuregulin 1 (NRG1) is one of the plausible hypotheses for the pathogenesis of schizophrenia. The neuregulin 1 (NRG1) is located on chromosome 8p, as suggested by multiple linkage studies. The aim of this study is to clarify the contribution of polymorphisms of the neuregulin 1 (NRG1) with schizophreniaMethodsAfter informed consent was obtained, 100 schizophrenia patients and 100 control subjects were enrolled in this study. All subjects were administered the Diagnostic Interview for Genetic Studies (DIGS) (National Institute of Mental Health-Molecular Genetics Initiative, 1992; Nurnberger et al., 1994) by a research assistant with extensive training in this interview. Blood samples were collected in anonymously identified 10-ml Vacutainer tubes (Becton Dickinson). DNA was prepared by a modified SDS/Proteinase K procedure (Gusells et al., 1979). We genotyped polymorphism neuregulin 1 (NRG1) with the PCR-RFLP methods. The PCR products were digested by restricted enzyme.ResultsWe observed a significant association between the polymorphism neuregulin 1 (NRG1) and the schizophrenia (Chi-Square Test P= 0.0449).DiscussionThe NRG1gene was originally identified as a susceptibility gene for schizophrenia by using a combination of a linkage and association approaches based on microsatellite markers and then using SNPs after microsatellite at risk haplotypes were identified. We found there is the frequency of the polymorphism of neuregulin 1 (NRG1) was significantly increased in schizophrenia patients. This allelic association suggests that the functional polymorphism neuregulin 1 (NRG1) may play a role in susceptibility to schizophrenia. Further study with larger sample sizes is required.

Creation of a Universal Sample Bank for Determining the 99th Percentile for Cardiac Troponin Assays.

International guidelines authored and endorsed by professional societies of cardiology, emergency medicine, and laboratory medicine are unanimous that the cutoff concentration for establishing a diagnosis of acute myocardial infarction be set at the 99th percentile of a healthy population. The establishment of the actual 99th percentile value is assay- and sample-dependent and is influenced by the population of individuals selected for testing. We created a sample bank that will enable manufacturers of troponin assays a consistent comparison of the 99th percentile. Participants were recruited from those attending the 2015 Annual Meeting of the AACC for the creation of a universal sample bank of apparently healthy individuals (free from uncontrolled diabetes, renal insufficiency, and heart disease). For those who met eligibility criteria and signed a written consent to participate, 60 mL blood was collected into 6 10-mL tubes each (2 serum, 3 heparin plasma, and 1 EDTA). Whole blood was tested for hemoglobin (Hb) A1c, and serum was tested for N-terminal pro-B-type natriuretic peptide (NT-proBNP) and creatinine. There were a total of 764 individuals who consented during the AACC Annual Meeting. After this initial enrollment, it was determined that there was an insufficient number of male participants recruited. Under the same protocol and consent, blood from 131 additional males was collected at the University of Maryland. Samples were centrifuged, and 240 μL aliquots of the 2 serum, 3 heparin plasma, and 1 EDTA tubes were frozen at 70 °C within 2 h of collection. The labeled samples were divided into boxes containing 1 aliquot from each individual. Sets of these samples were made available for purchase to manufacturers of cardiac troponin assays. Eighty-eight samples were excluded from the database for having a high NT-proBNP (>300 ng/L), low estimated glomerular filtration rate (eGFR) (<60 mL/min/1.73 m2), high Hb A1c (≥6.5%), or preanalytical sample issues and consenting/data issues. The final total was 808 individuals (402 females and 406 males; 60% Caucasian, 26% African or African American, 11% Asian or Pacific Islander, and 3% other). The creation of a bank of samples from healthy individuals enables a consistent comparison of the 99th percentile results from manufacturers of cardiac troponin assays.

Genetic polymorphisms to predict progression-free survival in patients with metastatic castration-resistant prostate cancer (mCRPC) receiving abiraterone therapy: Results from the NCI 9012 trial.

145 Background: Abiraterone is a CYP17 inhibitor approved for treatment of men with mCRPC. The NCI 9012 trial evaluated abiraterone alone with or without the PARP inhibitor veliparib in mCRPC patients. We hypothesized that germline genetic variation in the androgen axis and other metabolic enzymes would predict response to veliparib + abiraterone vs. abiraterone alone. Methods: A randomized trial cohort of (148) men with advanced mCRPC treated with abiraterone with or without veliparib was genotyped for 120 DNA polymorphisms in genes involved in androgen metabolism using Lifetech Open array chips. Blood for pharmacogenomic SNP analysis were collected at pre-treatment from each subject into 10-mL EDTA tube. Polymorphisms were tested using Cox models without treatment for prognostic testing and with treatment arm for predictive testing. Results: Genotyping was completed in 143 of 148 men; all were treated with abiraterone; 72 without veliparib (Median PFS: 10.3m) and 71 with veliparib (Median PFS: 11.3m). Polymorphisms in separate genes (SLCO2B1, KIF3C CYP19A, ESR1) were significantly (P ≤ .025) associated with progression-free survival (PFS) during abiraterone (q-value &lt; 0.69). Polymorphisms in (CYP11A1, HSD17B4, ABHD13;LIG4, CYP19A1, HSD17B4, TRMT11) were predictive for PFS in patients treated with combination of abiraterone/veliparib compared to abiraterone alone (p-value &lt; 0.025; q-value &lt; 0.28). Conclusions: This analysis examines the influence of inherited variations on the efficacy of abiraterone, establishing the importance of pharmacogenomics on individual’s response to this therapy. Genotyping patients at these loci could be predictive of improved PFS to valiparib in combination with abiraterone. Further analysis of the association of more than one polymorphisms compared to zero or one with PFS associated with improved TTP demonstrated a better response to therapy than individuals carrying zero or one is ongoing. Clinical trial information: NCT01576172.

The 9-MilCA method as a rapid, partly automated protocol for simultaneously recording milk coagulation, curd firming, syneresis, cheese yield, and curd nutrients recovery or whey loss

The aim of this study was to propose and test a new laboratory cheesemaking procedure [9-mL milk cheesemaking assessment (9-MilCA)], which records 15 traits related to milk coagulation, curd firming, syneresis, cheese yield, and curd nutrients recovery or whey loss. This procedure involves instruments found in many laboratories (i.e., heaters and lacto-dynamographs), with an easy modification of the sample rack for the insertion of 10-mL glass tubes. Four trials were carried out to test the 9-MilCA procedure. The first trial compared 8 coagulation and curd firming traits obtained using regular or modified sample racks to process milk samples from 60 cows belonging to 5 breeds and 3 farms (480 tests). The obtained patterns exhibited significant but irrelevant between-procedure differences, with better repeatability seen for 9-MilCA. The second trial tested the reproducibility and repeatability of the 7 cheesemaking traits obtained using the 9-MilCA procedure on individual samples from 60 cows tested in duplicate in 2 instruments (232 tests). The method yielded very repeatable outcomes for all 7 tested cheese yield and nutrient recovery traits (repeatability >98%), with the exception of the fresh cheese yield (84%), which was affected by the lower repeatability (67%) of the water retained in the curd. In the third trial (96 tests), we found that using centrifugation in place of curd cooking and draining (as adopted in several published studies) reduced the efficiency of whey separation, overestimated all traits, and worsened the repeatability. The fourth trial compared 9-MilCA with a more complex model cheese-manufacturing process that mimics industry practices, using 1,500-mL milk samples (72 cows, 216 tests). The average results obtained from 9-MilCA were similar to those obtained from the model cheeses, with between-method correlations ranging from 78 to 99%, except for the water retained in the curd (r=54%). Our results indicate that new 9-MilCA method is a powerful research tool that allows the rapid, inexpensive, and partly automated analysis processing 40 samples per day with 2 replicates each, using 1 lacto-dynamograph, 2 heaters, and 3 modified sample racks, and yields a complete picture of the cheesemaking process (e.g., milk gelation, curd firming, syneresis, and whey expulsion) as well as the cheese yield and the efficiency of energy or nutrients retention in the cheese or loss in the whey.

Effect of selenium and vitamin E supplementation on reproductive indices and biochemical metabolites in Raieni goats

The aim of this study was to investigate the effect of selenium (Se) and vitamin (vit) E supplementation on reproductive indices and biochemical metabolites in Raieni goats. Ninety adult Raieni goats, weighing 43 ± 2.5 kg and with a body condition score of 3.5 ± 0.1, were randomly allocated into nine groups which entered into an experiment with 3 × 3 factorial design. The experimental diets consist of three levels of se (0.0, 0.3 and 0.5 mg/kg of feed) and three levels of vit E (0.0, 20 and 50 mg/kg of feed). At the end of the experiment, blood samples were collected from the jugular vein of all goats into 10-ml vacuum tubes (venoject). Serum was recovered by centrifugation (10 m at 3000 rpm) and stored at –20°C until assayed for the concentration of biochemical metabolites (glucose, cholesterol, triglycerides, low-density lipoprotein (LDL), high-density lipoprotein (HDL), total proteins, albumin and T3 and T4). The results of this experiment revealed that supplementation of diets with Se or vit E had no significant effect on biochemical metabolites or reproductive traits. However, the interactive effect of Se and vit E on reproductive indices was significant (P < 0.05), with goats fed diets containing 0.5 mg/kg Se and 20 mg/kg vit E, giving birth to heavier kids compared to those which received the other dietary treatments.

Selective decomposition of proteins by photocatalytic Ti(IV)-doped calcium hydroxyapatite particles from mixed-protein systems

The decomposition of protein molecules from a mixed-protein solution on the surface of calcium hydroxyapatite (CaHap) and Ti(IV)-doped CaHap (TiHap) particles with a Ti/(Ca + Ti) atomic ratio (X Ti) of 0.10 and 0.20 under UV irradiation of 365 nm in wavelength was investigated. Acidic bovine serum albumin (BSA) and basic lysozyme (LSZ) were employed as a model of pathogenic proteins. The photocatalytic activities of TiHap particles were estimated from the decomposition of BSA and LSZ from the BSA (2.5 mg/cm3)–LSZ(1.0 mg/cm3) mixture under 1 mW/cm2 UV irradiation dispersed in a 10-mL quartz tube. No change in BSA concentration by UV irradiation was observed for all the unheated original CaHap and TiHap particles without and with low photocatalytic activities, respectively. Similar results were observed for the systems that employed heat-treated particles endowed a high photocatalytic activity by heat treatment at 650 °C for 1 h. On the other hand, a selective photocatalytic decomposition was observed for the LSZ, i.e., only LSZ molecules were decomposed completely from the BSA (2.5 mg/cm3)–LSZ(1.0 mg/cm3) mixture by using heat-treated TiHap particles with X Ti = 0.10 and 0.20. This selective decomposition by TiHap particles was interpreted by higher adsorption affinity of positively charged LSZ to highly negatively charged TiHap together with low molecular weight and rigid structure of LSZ molecules.

Abstract A28: Latino farm workers and non-farm workers and their experiences in providing biospecimens for research.

Abstract Objectives: The goal of this project was to re-contact 40 of the 100 adult participants (Hispanic farm worker and non-farm workers), who participated in a pesticides exposure study, to learn about their experience providing biospecimens for research. The 40 randomly selected participants are members of a larger cohort that has been participating in pesticides exposure research for over 10 years. During their participation in the pesticides exposure study these participants were asked to provide two 10mL tubes of blood, two buccal cell samples and four urine samples during the seven-day collection period. Two dust samples were also collected from the home and a 7-day dust sample was collected from the vehicle. This protocol was repeated a total of three times (during two different pesticide spray seasons and one non-spray season) over the course of one year. Twelve of the 40 participants interviewed also provided 20 saliva samples, 2 hair samples and one blood sample over 10 weeks for a stress and cortisol study. Methods: Two months after completing the pesticides exposure study, we re-contacted a random selection of 40 participants to take part in a one-on-one interview. All participants are Hispanic farm workers and non-farm workers and all live in the Yakima Valley of Washington. Interview questions were selected to learn about the participant's feelings toward research, the barriers and facilitators to participating in research and providing biological samples, expectations surrounding their biological samples and thoughts about providing tumor samples. Results: A total of 42 participants were re-contacted; 40 agreed to participate and completed an interview, 1 refused and 1 was lost to follow-up. Each participant who completed an interview was provided a $20 gift card. Qualitative analysis is summarizing the beliefs and opinions of Hispanics about donating biospecimens and their understanding of what such donation means. Their attitudes to research are also being explored. Results will be available in mid-July, 2012. Conclusion: Speculation about Latinos giving biospecimens suggests that Hispanics are more reluctant to participate than non-Hispanic Whites. This was not our experience. The personal meaning of donation and the meaning of research is important to better understand Latinos participation in biobanking. Citation Format: Beti Thompson, Elizabeth Carosso, Ilda Islas, Genoveva Ibarra. Latino farm workers and non-farm workers and their experiences in providing biospecimens for research. [abstract]. In: Proceedings of the Fifth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2012 Oct 27-30; San Diego, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2012;21(10 Suppl):Abstract nr A28.

Clinical transplantation ofex vivoexpanded autologous limbal epithelial cells using a culture medium with human serum as single supplement: a retrospective case series

Presently, our clinic is the only centre in Scandinavia that offers patients with corneal surface pathology including limbal stem cell deficiency (LSCD) transplantation of ex vivo expanded limbal epithelial cells (LECs). We here present clinical data of the first nine patients with LSCD who were transplanted with autologous LECs expanded in medium completely free of any animal-derived products and non-human/recombinant growth factors (including Cholera Toxin), and with autologous human serum as the only growth supplement. We conducted a noncomparative retrospective study of patients with LSCD at our centre between 2009 and 2011. The diagnosis was based on history and clinical signs. A biopsy was taken from healthy limbus, and the epithelium was expanded on amniotic membrane (AM) in medium containing autologous serum and subsequently transplanted to the affected eye. Successful outcome was defined as relief of pain and photophobia and/or improved best corrected visual acuity (BCVA) and/or reestablishment of a stable corneal epithelium and regression of corneal vascularization. Five of the nine transplanted patients (55.6%) had an improvement in either subjective symptoms or objective findings (11- to 28-month follow-up). Our clinical study shows that patients with LSCD can be treated successfully with transplantation of LECs expanded ex vivo in a medium with autologous serum as the only growth supplement. The use of this novel culture system, which is devoid of animal-derived products and non-human/recombinant growth factors (including Cholera Toxin), reduces the risks of inter-species disease transmission and host immune responses to xenogenic proteins, both obvious advantages for the patient.

Pharmacodynamic Effects of Simulated Standard Doses of Antifungal Drugs against Aspergillus Species in a New In Vitro Pharmacokinetic/Pharmacodynamic Model

In conventional ΜΙC tests, fungi are exposed to constant drug concentrations, whereas in vivo, fungi are exposed to changing drug concentrations. Therefore, we developed a new in vitro pharmacokinetic/pharmacodynamic model where human plasma pharmacokinetics of standard doses of 1 mg/kg amphotericin B, 4 mg/kg voriconazole, and 1 mg/kg caspofungin were simulated and their pharmacodynamic characteristics were determined against three clinical isolates of Aspergillus fumigatus, Aspergillus flavus, and Aspergillus terreus with identical MICs (1 mg/liter for amphotericin B, 0.5 mg/liter for voriconazole) and minimum effective concentrations (0.5 mg/liter for caspofungin). This new model consists of an internal compartment (a 10-ml dialysis tube made out of a semipermeable cellulose membrane allowing the free diffusion of antifungals but not galactomannan) inoculated with Aspergillus conidia and placed inside an external compartment (a 700-ml glass beaker) whose content is diluted after the addition of antifungal drugs by a peristaltic pump at the same rate as the clearance of the antifungal drugs in human plasma. Fungal growth was assessed by galactomannan production. Despite demonstrating the same MICs, amphotericin B completely inhibited (100%) A. fumigatus but not A. flavus and A. terreus, whose growth was delayed for 7.53 and 22.8 h, respectively. Voriconazole partially inhibited A. fumigatus (49.5%) and Α. flavus (27.9%) but not Α. terreus; it delayed their growth by 3.99 h (A. fumigatus) and 5.37 h (Α. terreus). Caspofungin did not alter galactomannan production in all of the species but A. terreus. The new model simulated human pharmacokinetics of antifungal drugs and revealed important pharmacodynamic differences in their activity.

Analysis of branched-chain α-keto acid dehydrogenase complex activity in rat tissues using α-keto[1- 13C]isocaproate as substrate

We have developed a highly sensitive nonradioisotope method for the measurement of branched-chain α-keto acid dehydrogenase complex (BCKDC) activity using α-keto[1- 13C]isocaproate ([1- 13C]KIC) as substrate. The enzyme reaction was performed in a 10-ml test tube. After incubation of the reaction mixture (1 ml), the reaction was terminated by acidification, followed by the addition of internal standard (3.5 μmol K 2CO 3) for calculation of 13CO 2. The 13CO 2 released to the gas phase in the test tube was calculated from the molar ratio of 13CO 2/ 12CO 2 assayed by gas chromatography isotope ratio mass spectrometry. A distinct linear relationship between 13CO 2 production rate and BCKDC activity was obtained when using purified BCKDC. The detection limit of 13CO 2 in the assay method was at least 0.167 nmol, which is similar to that in the previously reported radiochemical method. The enzyme assay was found to be linear with respect to reaction time and quantity of enzyme used up to approximately 60 nmol 13CO 2 produced. We measured BCKDC activities in several rat tissues, including skeletal muscle. The activities of BCKDC obtained are comparable to those reported previously. These results suggest that the novel BCKDC assay method using [1- 13C]KIC is useful for measurement in tissues with low BCKDC activity such as skeletal muscle.

Obtaining Blood Samples From Peripheral Intravenous Catheters: Best Practice?

Obtaining Blood Samples From Peripheral Intravenous Catheters: Best Practice?

Single-layer centrifugation with Androcoll-E can be scaled up to allow large volumes of stallion ejaculate to be processed easily

The objective of the current study was to optimize the volumes of Androcoll-E and sperm sample used in various sizes of centrifuge tube to scale up single-layer centrifugation (SLC) for routine use in the field. Although sperm suspensions of equivalent quality were produced using Androcoll-E in small and large tubes, the sperm yield was much lower in the latter (P < 0.001). In contrast, in 200-mL tubes (XL), the yields were approximately 25% higher than those for the small tubes. An increased volume (4.5 mL) of extended ejaculate in small tubes (SLC-Inc) or 15 to 18 mL extended ejaculate on 15 mL of colloid of a reduced density, Androcoll-E-Large (SLC-Large), in 50-mL tubes were both found to give similar yields of motile spermatozoa as that of the SLC-Small method (SLC-Small, 49.7 ± 18.6%; SLC-Inc, 53.3 ± 17.1%; SLC-Large, 44.9 ± 18.3%) and were found to be equivalent in quality (motility: 88.0 ± 8.8%, 84.0 ± 3.5%, 90.0 ± 5.4%; normal morphology: 69.4 ± 17.0%, 69.4 ± 12.7%, 63.9 ± 15.6%; viability: 78 ± 16.7%, 83.8 ± 12.5%, 80.05 ± 14.6%; DNA fragmentation index: 14.7 ± 10.9%, 12.8 ± 8.1%, 11.6 ± 7.6%, respectively). The processing of an “average” stallion Equus caballus ejaculate in approximately twenty-seven 10-mL tubes (SLC-Inc) or eight 50-mL tubes (SLC-Large) is feasible, the latter being considered more practical for on-stud use.