A Lab-in-a-briefcase for rapid prostate specific antigen (PSA) screening from whole blood

We present a new concept for rapid and fully portable Prostate Specific Antigen (PSA) measurement, termed “Lab-in-a-Briefcase”, which integrates an affordable microfluidic ELISA platform utilising a melt-extruded fluoropolymer Micro Capillary Film (MCF) containing 10 bore, 200 μm internal diameter capillaries, a disposable multi-syringe aspirator (MSA) plus a sample tray pre-loaded with all required immunoassay reagents, and a portable film scanner for colorimetric signal digital quantitation. Each MSA can perform 10 replicate microfluidic immunoassays on 8 samples, allowing 80measurements to be made in less than 15 minutes based on semi-automated operation and norequirement of additional fluid handling equipment. An assay was optimised for measurement of a clinically relevant range of PSA from 0.9 to 60.0 ng/ml in 15 minutes with CVs in the order of 5% based on intra-assay variability when read using a consumer flatbed film scanner. The PSA assay performance in the MSA remained robust in the presence of undiluted or 1:2 diluted human serum or whole blood, and the matrix effect could simply be overcome by extending sample incubation times. The PSA "Lab-in-a-briefcase" is particularly suited to a low-resource health setting where diagnostic labs and automated immunoassay systems are not accessible, by allowing PSA measurement outside the laboratory using affordable equipment.

An operationally simple sonogashira reaction for an undergraduate organic chemistry laboratory class

An operationally simple, reliable, and cheap Sonogashira reaction suitable for an undergraduate laboratory class that can be completed within a day-long (8 h) laboratory session has been developed. Cross-coupling is carried out between 2-methyl-3-butyn-2-ol and various aryl iodides using catalytic amounts of bis-(triphenylphosphine)palladium(II) dichloride, with copper(I) iodide as a cocatalyst, in triethylamine at room temperature, so a range of products can be prepared within a single group and results compared. The coupling itself is usually complete within 1.5 h and is easily monitored by TLC, leaving up to 6 h for purification and characterization. Purification is by “mini flash column chromatography” through a plug of silica encased in the barrel of a plastic syringe, so the procedure is amenable to large class sizes.

New-on-line method for water isotope analysis of speleothem fluid inclusions using laser absorption spectroscopy (WS-CRDS)

A new online method to analyse water isotopes of speleothem fluid inclusions using a wavelength scanned cavity ring down spectroscopy (WS-CRDS) instrument is presented. This novel technique allows us simultaneously to measure hydrogen and oxygen isotopes for a released aliquot of water. To do so, we designed a new simple line that allows the online water extraction and isotope analysis of speleothem samples. The specificity of the method lies in the fact that fluid inclusions release is made on a standard water background, which mainly improves the δ D robustness. To saturate the line, a peristaltic pump continuously injects standard water into the line that is permanently heated to 140 °C and flushed with dry nitrogen gas. This permits instantaneous and complete vaporisation of the standard water, resulting in an artificial water background with well-known δ D and δ18O values. The speleothem sample is placed in a copper tube, attached to the line, and after system stabilisation it is crushed using a simple hydraulic device to liberate speleothem fluid inclusions water. The released water is carried by the nitrogen/standard water gas stream directly to a Picarro L1102-i for isotope determination. To test the accuracy and reproducibility of the line and to measure standard water during speleothem measurements, a syringe injection unit was added to the line. Peak evaluation is done similarly as in gas chromatography to obtain &delta D; and δ18O isotopic compositions of measured water aliquots. Precision is better than 1.5 ‰ for δ D and 0.4 ‰ for δ18O for water measurements for an extended range (−210 to 0 ‰ for δ D and −27 to 0 ‰ for δ18O) primarily dependent on the amount of water released from speleothem fluid inclusions and secondarily on the isotopic composition of the sample. The results show that WS-CRDS technology is suitable for speleothem fluid inclusion measurements and gives results that are comparable to the isotope ratio mass spectrometry (IRMS) technique.

Multiplexed femtomolar quantitation of human cytokines in a fluoropolymer microcapillary film

Sensitive quantitation of multiple cytokines can provide important diagnostic information during infection, inflammation and immunopathology. In this study sensitive immunoassay detection of human cytokines IL-1β, IL-6, IL-12p70 and TNFα is shown for singleplex and multiplex formats using a novel miniaturized ELISA platform. The platform uses a disposable plastic multi-syringe aspirator (MSA) integrating 8 disposable fluoropolymer microfluidic test strips, each containing an array of ten 200 mean i.d. microcapillaries coated with a set of monoclonal antibodies. Each MSA device thus performs 10 tests on 8 samples, delivering 80 measurements. Unprecedented levels of sensitivity were obtained with the novel fluoropolymer microfluidic material and simple colorimetric detection in a flatbed scanner. The limit of detection for singleplex detection ranged from 2.0 to 15.0 pg/ml, i.e. 35 and 713 femtomolar for singleplex cytokine detection, and the intra- and inter-assay coefficient of variation (CV) remained within 10%. In addition, a triplex immunoassay was developed for measuring IL-1β, IL-12p70 and TNFα simultaneously from a given sample in the pg/ml range. These assays permit high sensitivity measurement with rapid <15 min assay or detection from undiluted blood serum. The portability, speed and low-cost of this system are highly suited to point-of-care testing and field diagnostics applications.

Influência do tipo de parto na hematologia do cordão umbilical de cães

O sangue do cordão umbilical humano tem sido crescentemente utilizado como fonte de células-tronco. Os modelos experimentais de células-tronco da medula óssea, em cães, têm propiciado informações importantes para transplantes medulares em humanos. Vários trabalhos citam a influência do tipo de parto nas características do sangue do cordão umbilical (SCU) humano. Entretanto, não existem relatos sobre a colheita do sangue do cordão umbilical de cães. O objetivo deste ensaio é avaliar a influência do tipo de parto na hematologia do cordão umbilical de cães. No presente protocolo experimental, foram estudados 54 fetos de cães, ao final da vida intra-uterina, provenientes de parto normal (n=24) e cesariana (n=30). A colheita de sangue do cordão umbilical foi realizada com seringa de cinco mL contendo solução anticoagulante EDTA (1mg/1mL sangue). em seguida, a contagem global de hemácias, leucócitos, plaquetas, a determinação da concentração de hemoglobina, taxa de hematócrito, os índices eritrocitários foram realizados no contador automático de células. A contagem diferencial de leucócitos foi determinada em esfregaços de SCU corados com May-Grunwald-Giemsa (MGG). Com relação ao eritrograma e ao plaquetograma, não houve diferença significativa entre as amostras obtidas em cesarianas e partos normais. Os valores do leucograma do SCU colhido em partos normais foram superiores àqueles obtidos em cesarianas (P<0,05). Portanto, o tipo de parto influencia os valores hematológicos do cordão umbilical de cães.

Uma intervenção educativa para profissionais de saúde na prevenção de pneumonia associada à ventilação mecânica

Quasi-experimental study, prospective with quantitative approach, performed at the Hospital do Coração in Natal, aimed at verified the existence of difference between the care given by health professionals to the patients under mechanical ventilation (MV) in the Intensive Care Unit, before and after an educative intervention. The population was of 31 professionals, with data collected between november 05 of 2007 to march 27 of 2008. The results show a yong population, female gender, middle level of education, nursing technique, working between 05 and 09 years on nursing profession, and 01 to 04 years on Intensive Care Unit; almost all, never had an kind of training over prevent pneumonia associated to mechanical ventilation; from those that had training, occur on the work place with duration from 12 to 24 hours. About endotracheal intubation, the cuff was tested with a sterilized syringe had a positive change after a educative intervention, increased from 75,0% to 100,0%; the sterile guide was used on 75,0% before and 100,0% after an educative intervention. Regarding endotracheal suction procedure, was not informed to the patient on 72,7% before, however was informed on 56,7% after; the hands was not previously washed 68,5% before, however was 63,3% after the procedure; mask was used on 74,2 % opportunities before and 76,7% after; the aspiration catheter had adequated size on 98,9% observation before and 100,0% after; the gaze was sterilized on 95,7% before and 100,0% after; the ventilator was connected to the patient during the aspiration intervals on 94,4% observation before and 100,0% after; the ambu bag was clean and protected on 76,1% before and 85,7% after; the aspiration catheter was discarded after be used on 98,9% before and 100,0% after; FIO2 was turned to the begging value on 32,9% observation before and 12,0% after; before the procedure 71,9% professions washed their hands and 73,3% after; before, notes of aspiration results were performed on 70,8% observation and 86,7% after. Regarding devices used on respiratory tract, aspirator flasks were not swapped on 84,6% observations before and 71,0% after; daily látex extention change was not performed on 93,6% observation before and 87,1% after; the ambu bag change was not performed on 50,0% observation before even if was duty or unprotected and on 75,8% opportunities was changed, after; nebulization was not prepared with sterile fluids or manipulated aseptically on 65,2% observation before, perhaps was on 71,7% after; before nebulizers were not changed on 65,2% observations, perhaps were on 60,9% after. Concerning ventilator breathing circuits, condense fluids cumulated on circuits were removed on 55,0% opportunities before, and 64,0% after; moisturizer was not filled with sterile water when already had small amount of liquid inside on 78,4% observations before, and 90,2% after; MV circuits were changed on 97,0% observations on presence of visible duty or when presents some kind of failure, before and 98,4% after. About body position, on 51,3% observations the decubitus position change were done before and 78,2% after; fowler position was maitened on 95,5% observations before and 98,2% after; Regarding respiratory physiotherapy, enteral diet was not interrupted before respiratory physiotherapy on 94,9% before and 90,0% after; respiratory physiotherapy devices were not disinfected or sterile on 69,6% observations before but they re on 60,0% after; before the cateter was not tested before introduction enteral diet or medications on 100,0% but after was done on 15,2%. About enteral feeding, intestine motility and measure of stomach contents were not done on 100,0% observations before, but was 15,2% after. We conclude that 05 of 07 valuated procedures in relation to MV, had a significant improvement on quality of care given after educative intervention, when compared before intervention

Comparative study between target-controlled-infusion and continuous-infusion anesthesia in dogs treated with methotrimeprazine and treated with propofol and remifentanil

OBJETIVO: Comparar duas técnicas de infusão de propofol em cadelas submetidas à ovariohisterectomia, estudando a eficácia da dose alvo de propofol, avaliando os atributos cardiorrespiratórios, hemogasométricos e escala do índice bispectral, (BIS) bem como as características do período de recuperação. MÉTODOS: Foram anestesiadas 20 cadelas, distribuídos em dois grupos (GI e GII). em GI, os animais foram pré-tratados com levomepromazina e anestesiados com propofol por infusão alvo controlada, através de bomba de infusão Harvard pump, associado com remifentanila, através de bomba de seringa. em GII, os animais receberam o mesmo tratamento de GI, só que ao invés de receberem o propofol por infusão alvo controlada, receberam o propofol em infusão contínua de velocidade fixa. RESULTADOS: Bradicardia e discreta hipotensão, estabilidade hemogasométrica e respiratória, além de um bom miorrelaxamento, mais evidente na infusão contínua e boa hipnose. CONCLUSÕES: As doses de propofol utilizadas em ambas as técnicas, após o pré-tratamento de levomepromazina e associadas ao opióide, foram eficazes para a realização cirúrgica. A técnica de anestesia alvo controlada obteve um menor consumo de anestésico (propofol) com períodos mais rápidos de recuperação.

Improvement in embryo recovery using double uterine flushing

The objective of the present study was to evaluate the effects of double uterine flushing on the recovery of embryos/ova in cattle. Two hundred and ten embryo recovery procedures were conducted using a double uterine flushing method, and the results were compared with 432 conventional single-flushing procedures. Cyclic Limousin (n = 403) and Guzera (n = 239) donor cows received an intravaginal progesterone releasing device and 2 mg of estradiol benzoate on Day 0. Between Days 5 and 9, donors received decreasing doses of FSH, which ranged from 200 to 300 IU (Bos indicus) and 300 to 500 IU (Bos taut-us). on the afternoon of Day 7, donors received an injection of 500 mu g cloprostenol and progesterone implants were removed 12 It later (morning of Day 8). Artificial insemination was performed between 14 and 26 h after first detection of behavioral estrus. Cows were randomly assigned to have embryos recovered by a double-flushing method (n = 210) or the conventional single-flushing procedure (n = 432). For the double-flushing procedure, after first flushing the whole uterus with 1 L of Dubelco's Phosphate Buffered Saline (DPBS), a Foley catheter was positioned in the uterine body to permit refilling of the uterus with fresh DPBS (80-150 mL). The catheter was closed with the plunger of a disposable 5 mL syringe, and the donors were allowed to rest in a holding area for 30 min. Thereafter, a second flush was performed to recover the solution remaining in the uterus. Animals from the control group were subjected to a single uterine flush. From 2 10 double-flushing procedures, 1409 viable embryos were recovered. In comparison, from 432 cows receiving the single-flushing procedure, 1993 embryos were recovered. Double flushing increased (P < 0.05) the number of embryos recovered per procedure compared to single flushing (6.7 +/- 0.4 versus 4.6 +/- 0.2, respectively; mean +/- S.E.M.). When double flushing was performed, average recovered embryos/ova increased (P < 0.05) from 8.3 +/- 0.4 to 12.7 +/- 0.7 in Limousin and from 7.9 to 11.5 in Guzera. Also, utilization of double flushing increased (P < 0.05) the number of viable embryos from 4.7 +/- 0.3 to 6.9 +/- 0.5 in Limousin and from 4.5 +/- 0.4 to 6.4 +/- 0.7 in Guzera. Mean total embryos/ova was similar (P > 0.05) between the control group and after the first uterine flushing in the double-flushing group; therefore, both flushings were conducted efficiently. In conclusion, double uterine flushing increased embryo recovery in cattle. (C) 2004 Elsevier B.V. All rights reserved.

Comparison of the biocompatibility of different root canal irrigants

The purpose of this study was to compare the reaction of rat subcutaneous connective tissue to 0.9% sterile saline, 2.5% sodium hypochlorite (NaOCl), 5.25% NaOCl and 2% chlorhexidine gluconate solution or gel. Six circles were demarcated on the dorsal skin of 24 male Wistar rats, leaving 2 cm between each circle. Using a syringe, 0.1 mL of each root canal irrigant was injected subcutaneously into 5 circles. In the 6th circle, the needle of an empty syringe was introduced into the skin, but no irrigant was injected (control group). Evaluations were undertaken at 2 h, 48 h, 14 days and 30 days post-procedure. Tissue samples were excised, embedded in paraffin blocks and 3-mu m-thick sections were obtained and stained with hematoxylin and eosin. The areas of inflammatory reaction were evaluated and analyzed statistically by ANOVA and Tukey's test. The control group showed few or no inflammatory reaction areas in the subcutaneous tissue. 0.9% saline solution, 2.0% chlorhexidine solution and 2.5% NaOCl showed a good biocompatibility, as very mild inflammatory reaction was detected at 14 days and tissue repair occurred at 30 days. 5.25% NaOCl was the most toxic irrigant, as the number of inflammatory cells remained elevated at 14 and 30 days. The group treated with 2.0% chlorhexidine gluconate gel presented a moderate inflammatory response at 14 days, which decreased at 30 days, being considered similar to that of the control group, 0.9% saline solution, 2.0% chlorhexidine solution and 2.5% NaOCl at this experimental period.

Influences of the insertion method in glass ionomer cement porosity

The aim of this study was to evaluate the presence of porosities inside the glass ionomer cement (GIC) after different techniques of material insertion. MATERIAL and METHOD: Specimens were prepared with high-viscosity GIC Ketac Molar Easymix and divided into three groups according to the insertion method: spatula (PI), Centrix injector (CI), and low-cost syringe (LCS). The specimens were fractured and observed with scanning electronic microscopy to quantitatively evaluate porosity inside the material using Image J Software. RESULTS: Statistical analysis, ANOVA application, and Tukey test to significance level of 5%, revealed that there was no statistical difference between the groups. CONCLUSION: Although the use of LCS has not decreased the porosity of the material, this insertion method is easy, accessible, and low cost, which makes it a viable alternative of use in the ART technique and in others bucal health programs. Microsc. Res. Tech., 2012. (c) 2012 Wiley Periodicals, Inc.

Suscetibilidade de operárias e larvas de abelhas sociais em relação à ricinina

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Avaliação algimétrica por estímulo nociceptivo térmico e pressórico em cães pré-tratados com levomepromazina, midazolam e quetamina associados ou não ao butorfanol ou buprenorfina

OBJETIVO: Quantificar a dor em cães sob anestesia dissociativa através de estímulo térmico e pressórico e o período hábil de dois analgésicos opióides. MÉTODOS: Empregaram-se 30 cães alocados em três grupos (n=10), onde os animais de GI receberam levomepromazina e midazolam associado na mesma seringa à quetamina. Os animais de GII receberam o mesmo tratamento de GI porém associado ao butorfanol e por fim os animais de GIII receberam o mesmo tratamento de GI associando-se a buprenorfina. Procedeu se a avaliação paramétrica rotineira, empregando-se, entretanto, a termoalgimetria mensurada em °C em média de 52°C e a pressoalgimetria em kg. RESULTADOS: Na termoalgimetria houve diferença significativa em GI nos momentos M0 e M1,e em M4 e M5. em GII houve diferença em M0, M1, M5 e M6. em GIII houve diferença entre momentos M0 e M1. Na pressoalgimetria houveram diferenças em GI em diferentes momentos: M0, M2 e M3. em GII observaram-se diferenças em todos os momentos. em GIII observaram-se diferenças em M0 e M9. Ocorreram diferenças entre os grupos, sendo o M2 de GII menor que GI e GIII. Já em M4 e M5 de GIII demonstrou-se maior que GI e GII. E na avaliação dos períodos observou-se um período de latência significativamente maior em GI, porém com período hábil e de recuperação menor em relação à GII e GIII. Já a recuperação do tônus postural foi maior em GIII seguido de GII e finalmente de GI. CONCLUSÃO: O método empregado para mensuração do estímulo álgico foi eficiente, observando-se um período hábil analgésico de 3 horas para o butorfanol e de 6 horas para a buprenorfina.

Hemi-ovariossalpingohisterectomia em pacas prenhes e posterior ocorrência de prenhez (Agouti paca, Linnaeus, 1766)

Foi descrita a hemi-ovariossalpingohisterectomia em cinco pacas prenhes mantidas em cativeiro no Setor de Animais Silvestres da Faculdade de Ciências Agrárias e Veterinárias (FCAV-UNESP) de Jaboticabal, São Paulo, Brasil, a fim de observar futura prenhez no corno restante. A tranqüilização foi obtida após aplicação de azaperone (4mg/kg) seguida da aplicação de sulfato de atropina (0,06mg/kg) e da associação de cloridrato de quetamina (20mg/kg) e cloridrato de xilazina (1,5mg/kg), ambos na mesma seringa, para indução da anestesia. A anestesia geral foi obtida mediante inalação de halotano por máscara. Por meio de laparotomia mediana, foram retirados o corno uterino prenhe, o ovário e a tuba uterina, todos do mesmo antímero. Antibióticos (30.000UI/kg de três penicilinas e 12,5mg/kg de duas estreptomicinas) e analgésico (0,02mg/kg de buprenorfina) foram aplicados imediatamente após a cirurgia, sendo repetidos após dois dias. Todas as aplicações foram feitas por via intramuscular. Apesar da permanência de apenas um ovário após a cirurgia, nova prenhez ocorreu no corno restante nas cinco fêmeas submetidas à cirurgia, com o nascimento de filhotes (apenas um por parto) após 215, 248, 276, 302 e 310 dias da hemi-ovariossalpingohisterectomia.

Citologia aspirativa por agulha fina (CAAF), em testículo de onça pintada (Panthera onca), utilizada como ferramenta no diagnóstico de infertilidade

Técnicas de biópsia, caracterizadas pela remoção de segmentos de órgãos e tecidos para análise histopatológica, não são indicadas no auxílio diagnóstico de alterações testiculares para animais ameaçados de extinção, por não serem totalmente isentas de riscos. Neste sentido, é de grande interesse que se desenvolvam técnicas de biópsia testicular cada vez mais seguras e com o mínimo de conseqüências negativas. Com este intuito três onças pintadas (Panthera onca) foram submetidas a exames de Citologia Aspirativa por Agulha Fina (CAAF). Amostras foram obtidas através da punção aspirativa dos testículos, esfregaços foram confeccionados, corados com Panótico e analisados sob Microscopia Óptica. Simultaneamente foram realizadas coletas de sêmen para avaliação do volume, pH, concentração, motilidade, vigor e morfologia espermáticas. Quanto à avaliação espermática, os animais apresentaram valores semelhantes aos encontrados na literatura quanto ao volume, pH, motilidade, vigor e morfologia espermáticas. Quanto a concentração espermática os animais apresentaram valores abaixo dos encontrados na literatura. Nos exames de CAAF, todas as gerações de células germinativas foram identificadas, indicando espermatogênese normal em todos os animais, com exceção das espermátides finais duplas que ainda não foram relatadas como achados em punções testiculares de outras espécies, o que vem confirmar a elevada porcentagem de células teratológicas encontradas nesses animais. Desta forma, podemos concluir que a CAAF testicular é um método diagnóstico auxiliar importante na detecção de alterações testiculares em casos de sub ou infertilidade, podendo ser utilizados na rotina de investigação do trato reprodutivo masculino, quando o exame histopatológico, por ser um método altamente invasivo, é desaconselhável.

Characterization and quantification of blood cells from the umbilical cord of dogs

Background: Models for the study of hematopoietic stem cells in dogs provide important information for bone marrow transplantation in humans. Recent studies have reported the importance of human umbilical cord blood (UCB) as an alternative to allogenic bone marrow for hematopoietic reconstitution. However, there are no studies on the UCB cells of dogs. Objective: the aim of this experiment was to characterize and quantify the blood cells of the umbilical cord of dogs. Methods: the blood of the umbilical cord of 20 neonatal dogs, delivered at term, with a median gestation time of 58 days, was collected with a 5-mL syringe containing EDTA. Total RBC, WBC, and platelet counts, HCT, hemoglobin (Hgb) concentration, and RBC indices were determined using an automatic cell counter. The differential leukocyte count was determined manually in blood smears stained with May-Grunwald-Giemsa. Reticulocyte percentages were determined on blood smears stained with brilliant cresyl blue and counterstained with May-Grunwald Giemsa. Results: the MCHC and numbers of RBCs, WBCs, neutrophils, and eosinophils in UCB were lower as compared with reference values for the peripheral blood of healthy neonatal and adult dogs; whereas, the MCV and reticulocyte percentages were higher. Conclusion: Erythrocyte macrocytosis and hypochromasia in UCB were consistent with marked reticulocytosis and indicative of high erythropoietic activity. The results of this study are an important first step in the characterization of UCB from neonatal dogs.