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Background:. Limited long-term safety data are published on HA/CaHA/L, a hybrid dermal filler combining hyaluronic acid (HA), calcium hydroxyapatite (CaHA), and lidocaine (L). Methods:. This retrospective multicenter study assessed treatment-emergent adverse events (TEAEs) in adults treated with HA/CaHA/L. The full analysis set (FAS) included eligible consented adults (N = 403); the long-term safety analysis (LTSA) set included FAS participants with greater than or equal to 12-months HA/CaHA/L exposure (n = 243). Results:. Participants were majority female (94.0%), with Fitzpatrick skin phototypes II/III (80.1%) and a mean age of 50.1 years. Most participants (86.4%) received one HA/CaHA/L treatment. The median time between participants’ first HA/CaHA/L treatment and chart review was 15.4 months. Participants received a mean of 2.2 mL (0.5-8.9 mL) filler per treatment. Treated areas were predominantly malar (71.2%) and mandible (69.7%) regions. Most participants (95.0%) had one or more aesthetic treatments other than HA/CaHA/L [eg, other dermal fillers (84.1%), botulinum toxin (63.3%)]. Nineteen (4.7%) FAS participants had 20 documented TEAEs; most (3.5%, n = 14 participants) were mild in severity. Twelve TEAEs in 11 participants (2.7%) were related to HA/CaHA/L: induration (three, 0.7%), edema (3, 0.7%), and implant site nodules (five, 1.2%), which were noninflammatory and likely related to product placement. Among the LTSA, 15 (6.2%) participants had 16 documented TEAEs (six edema, five implant site nodules, one inflammation, three skin induration, one hypersensitivity); most were mild in severity. Nine TEAEs in eight participants (3.3%) were HA/CaHA/L-related. No treatment-emergent serious AEs were reported. Conclusion:. The data from this noninterventional retrospective study support the favorable longer term (>12 month) safety profile of HA/CaHA/L.

Background: This article describes a standardized, minimally invasive approach for harvesting the quadriceps tendon through a 2 to 3 cm transverse skin incision, presenting it as a viable option. Indications: This procedure is indicated for patients undergoing anterior cruciate ligament (ACL), posterior cruciate ligament, medial collateral ligament, or lateral collateral ligament reconstruction surgery using soft tissue quadriceps grafts. Technique Description: A transverse incision is made on the anterior face of the knee, followed by a longitudinal incision of the central third of the rectus femoris. We perform the distal detachment of the patella of the same tendon. Then, the tendon is extracted using a closed stripper. The graft is prepared and folded on itself, resulting in a double graft. Results: Twenty patients, 16 male and 4 female with complete rupture of ACL, were submitted to an ACL reconstruction using the quadriceps soft tissue graft with minimally invasive harvesting technique. The results were satisfactory for all patients. All patients showed good evolution in the immediate and late postoperative period, with no cases of joint stiffness, wound dehiscence, and no infections. Discussion/Conclusion: We can conclude that harvesting the quadriceps tendon with a minimally invasive approach is a valid and reliable option for the treatment of ligament tears of the knee. Patient Consent Disclosure Statement: The author(s) attests that consent has been obtained from any patient(s) appearing in this publication. If the individual may be identifiable, the author(s) has included a statement of release or other written form.

Background: Upgrade to cardiac resynchronization therapy (CRT) is common in Europe, despite little and conflicting evidence. Objective: To compare long-term clinical outcomes in a cohort of patients receiving de novo or upgrade to CRT. Methods: Single-center retrospective study of 295 consecutive patients submitted to CRT implantation between 2007 and 2018. Upgraded and de novo patients complying with a dedicated follow-up protocol were compared in terms of clinical (NYHA class improvement without major adverse cardiac events [MACE] in the first year of follow-up) and echocardiographic (left ventricle end-systolic volume reduction of >15% during the first year) response. Results: No differences in the rate of clinical (59.3% vs 62.6%, P = .765) or echocardiographic response (72.2% vs 71.9%, P = .970) between groups were observed. Device-related complications were also comparable between groups (8.9% vs 8.4%, P = .892). Occurrence of MACE and all-cause mortality were analyzed over a median follow-up of 3 (interquartile range 1–6) years: MACE occurred less frequently in the de novo group (hazard ratio [HR]: 0.55, 95% confidence interval [CI]: 0.34–0.90, P = .018), but all-cause mortality was similar among groups (HR: 0.87, 95% CI: 0.46–1.64, P = .684). Propensity score–matching analysis was performed to adjust for possible confounder variables. In the propensity-matched samples, all-cause mortality (HR: 1.26, 95% CI: 0.56–2.77, P = .557) and MACE (HR: 0.84, 95% CI: 0.46–1.54, P = .574) were comparable between upgrade and de novo patients. Conclusion: Survival after upgrade to resynchronization therapy was comparable to de novo implants. Additionally, clinical and echocardiographic response to CRT in upgraded patients were similar to de novo patients.

The aim of this study was to characterize the acute complications that required hospitalization in children under 5 years with Sickle Cell Disease. The population included 70 patients, 64% of African ascendency and 95.7% homozygotic for the HbS. We analyzed data from 337 hospitalizations. The first hospitalization occurred before 12 months in 38.6% and the average hospital admission per child was 3.92. The mean duration per hospitalization was 5.36 days. The most common diagnosis was vaso-occlusive crisis (36.8%) followed by fever, upper airway infections, bacterial pneumonia and splenic sequestration. Hemoglobin values at admission were between 6 and 7 g/dL in 22.3%, with 44.2% requiring at least 1 blood transfusion. There are several acute complications of SCD, being vaso-occlusive crisis the most common. Splenic sequestration generally occurs during the first 2 years and is associated with the need of transfusion. They represent a significant burden, with each child spending approximately 3 weeks hospitalized.

Objectives: The majority of available scores to assess mortality risk of coronavirus disease 2019 (COVID-19) patients in the emergency department have high risk of bias. Therefore, this cohort aimed to develop and validate a score at hospital admission for predicting in-hospital mortality in COVID-19 patients and to compare this score with other existing ones. Methods: Consecutive patients (≥ 18 years) with confirmed COVID-19 admitted to the participating hospitals were included. Logistic regression analysis was performed to develop a prediction model for in-hospital mortality, based on the 3978 patients admitted between March–July, 2020. The model was validated in the 1054 patients admitted during August–September, as well as in an external cohort of 474 Spanish patients. Results: Median (25–75th percentile) age of the model-derivation cohort was 60 (48–72) years, and in-hospital mortality was 20.3%. The validation cohorts had similar age distribution and in-hospital mortality. Seven significant variables were included in the risk score: age, blood urea nitrogen, number of comorbidities, C-reactive protein, SpO2/FiO2 ratio, platelet count, and heart rate. The model had high discriminatory value (AUROC 0.844, 95% CI 0.829–0.859), which was confirmed in the Brazilian (0.859 [95% CI 0.833–0.885]) and Spanish (0.894 [95% CI 0.870–0.919]) validation cohorts, and displayed better discrimination ability than other existing scores. It is implemented in a freely available online risk calculator (https://abc2sph.com/). Conclusions: An easy-to-use rapid scoring system based on characteristics of COVID-19 patients commonly available at hospital presentation was designed and validated for early stratification of in-hospital mortality risk of patients with COVID-19.

Background: Although biceps tenodesis has been widely used to treat its pathologies, few studies looked at the objective evaluation of elbow strength after this procedure. The purpose of this study is to clinically evaluate patients submitted to long head of the biceps (LHB) tenodesis with interference screws through an intra-articular approach and analyze the results of an isokinetic test to measure elbow flexion and forearm supination strengths. Methods: Patients who had biceps tenodesis were included in the study if they had a minimum follow-up of 24 months. Patients were excluded if they had concomitant irreparable cuff tears or previous or current contralateral shoulder pain or weakness. Postoperative evaluation was based on University of California–Los Angeles (UCLA) shoulder score and on measurements of elbow flexion and supination strength, using an isokinetic dynamometer. Tests were conducted in both arms, with velocity set at 60º/s with 5 concentric-concentric repetitions. Results: Thirty-three patients were included and the most common concomitant diagnosis were rotator cuff tear (69%) and superior labrum anterior to posterior (SLAP) lesions (28%). The average UCLA score improved from 15.1 preoperatively to 31.9 in the final follow-up (P < .001). Isokinetic tests showed no difference in peak torque between the upper limbs. One patient had residual pain in the biceps groove. None of the patients had Popeye deformity. UCLA score and follow-up length did not demonstrate correlation with peak torque. Conclusion: Arthroscopic proximal biceps tenodesis with interference screw, close to the articular margin, yielded good clinical results. Isokinetic tests revealed no difference to the contralateral side in peak torque for both supination and elbow flexion.

A 23-year-old man was admitted for acute pericarditis that evolved to cardiac tamponade and shock with need of emergent pericardiocentesis and inotropic support. Corticosteroid therapy was successful, but despite a gradual tapering, the patient relapsed. Incidentally, the patient developed hyperkalemia with hyponatremia. Subsequent hormonal measurements confirmed autoimmune polyglandular syndrome type-2. (Level of Difficulty: Intermediate.)

Background: Premature ejaculation (PE) prevalence can vary according to different definitions, assessment methods and populational demographics and culture. Aims: To investigate the differences between men classified as having “probable PE” (PEDT≥11), “possible PE” (PEDT = 9 or 10) or “no PE” (PEDT≤8) according to the Premature Ejaculation Diagnostic Tool (PEDT) criteria in regard to sociodemographic characteristics, and sexual and relational behavior. To assess the agreement of prevalence of PE according to 3 assessment methods: (i) the ejaculation latency time (ELT) according to the participant's memory; (ii) PEDT and (iii) a direct question about the self-perception of ejaculation as being normal, too early (premature) or retarded. Methods: In this web-based cross-sectional study, men aged ≥ 18 years living in the metropolitan region of São Paulo, Brazil, responded anonymously to an online survey. We used multinomial regression to estimate the association between PE according PEDT criteria and other features and the kappa coefficient to estimate agreement between the assessment methods. Outcomes: Association between PEDT-PE, sociodemographic characteristics and sexual and relational behaviors; agreement between PEDT, ELT and self-perception of PE. Results: Obesity, trying to hold back ejaculation, short or nonexistent foreplay and age

An 83-year-old man with significant background comorbidities was admitted with an inferior ST-segment elevation myocardial infarction. During primary percutaneous coronary intervention, a giant aneurysm is seen in the right coronary artery. (Level of Difficulty: Beginner.)

Infective celiac artery aneurysm is an extremely rare diagnosis, with few reported cases in the literature. We present the case of a rapidly expanding celiac artery aneurysm involving the ostia, probably infectious, successfully treated in an urgent setting by aneurysm exclusion resorting to the chimney technique. On follow-up, computed tomography angiography revealed complete aneurysm thrombosis and patent celiac artery. Previous reports of endovascular treatment of infective celiac artery aneurysm involved its embolization. This is the first reported case of chimney technique used to exclude a celiac artery aneurysm, with a clinical suspicion of infectious etiology, preserving celiac artery patency. Short-term results are encouraging, but implantation of prosthetic material in an infected environment is a concern. Keywords: Aneurysm, Infected, Celiac artery, Endovascular procedures

Congenital anomalies of the aorta are rare disorders that result from an abnormal development of the embryonic pharyngeal arch system. Aortic arch abnormalities occur in 1% to 2% of the population, and their clinical representation, usually in the first years of life, depend on esophageal or tracheobronchial compression or abnormal blood patterns. Such abnormalities are seldom seen in adults. (Level of Difficulty: Intermediate.)

Category: Midfoot/Forefoot, Trauma Introduction/Purpose: Acute avulsion fractures of the base of the fifth metatarsal are common and usually treated non- surgically with satisfactory outcomes. Forms of conservative treatments are described such as elasticated bandaging, immobilization in a cast, walking boot and hard-soled-shoes. The aim of this study was to compare the clinical and functional outcomes, time to return to the previous activities, time and rate of fracture healing of the patients with avulsion fracture of the base of the fifth metatarsal treated in a hard-soled-shoes (HSS) or in a walking boot (WB). Our null hypothesis is that a less rigid immobilization without restriction of the ankle yields the same final results. Methods: A retrospective and comparative study of 72 patients with acute avulsion fractures of the base of the fifth metatarsal treated with the WB or the HSS, from March 2014 to November 2018. The average age of the patients was 41,25 (range, 11-88) and there were 56 female and 16, male. 39 were treated with the WB and 33, with the HSS. Patients with comorbities that could interfere in the rate and time of bony healing (diabetes and inflammatory joint disease), associated ankle ligament lesions and loss of follow-up were excluded. Patients were followed regularly until they were clinically asymptomatic and able to return to their previous activities (work, daily activities, sports). We retrospectively reviewed medical records containing the visual analogue scale (VAS) for pain and the american orthopaedic foot and ankle society (AOFAS) score of each visit as well as radiographic exams to evaluate time and rate of fracture healing. Results: Patients of both groups had similar VAS and AOFAS scores in 8 (p=0,34 and p=0,83) and 12 (p=0,25 and p=0,79) weeks. Age and gender distribution in the two groups were equal (p=0,23 and p=0,34, respectively). Time taken to return to the previous activities were not significant different between both groups, with the HSS group taking 8,33 weeks and the WB taking 9,73 weeks (p=0,10). The mean time for bone consolidation was significant higher in the HSS group, taking 8,64 weeks while in the WB took 7,18 weeks (p

Introduction: Studies disagree as to whether there is a greater prevalence of hypertension among HIV/AIDS patients and the role of antiretroviral therapy. Objective: Evaluate the prevalence of hypertension and risk factors in a cohort of HIV-infected patients, with emphasis on antiretroviral therapy. Method: Case-control study conducted at baseline of a cohort, between June/2007 and December/2008 in Pernambuco/Brazil. Blood pressure was classified as normal, prehypertension, and hypertension. Results: Of 958 patients, 245 (25.6%) had hypertension (cases), 325 (33.9%) had prehypertension, and 388 (40.5%) were normotensive (controls). Comparison between hypertensive and normotensive patients showed that traditional factors, such as age > 40 (OR = 3.06, CI = 1.91- 4.97), male gender (OR = 1.85, CI = 1.15-3.01), BMI > 25 (OR = 5.51, CI = 3.36-9.17), and triglycerides > 150 mg/dL (OR = 1.69, CI = 1.05-2.71), were independently associated with hypertension. Duration of antiretroviral therapy and CD4 > 200 cells/mm3 were associated with hypertension in univariate analysis, but did not remain in final model. Type of antiretroviral schema and lipodystrophy showed no association with hypertension. Conclusion: Hypertension in HIV/AIDS patients is partially linked to invariable factors, such as age and sex. Efforts should be directed toward controlling reversible factors, particularly excessive weight gain and unsuitable diet. Keywords: HIV/AIDS, hypertension, antiretroviral therapy, cardiovascular risk

During last decade, male athletes were involved in official rhythmic gymnastics (RG) competitions. Aim of this study was to examine anthropometric characteristics and motor skills of male rhythmic gymnasts to identify guidelines in talent identification, selection and development training plans. Twenty-four gymnasts (age range 22+/-4 years), 12 male athletes and 12 female athletes, underwent two testing sessions: the previous session to assess anthropometric measures, and the second one to evaluate jumping ability using Optojump. Three vertical jumps: squat jump (SJ), counter-movement jump (CMJ), hopping test (HT) and three different technical jumps (Split Leap with stretched legs (SL); Cossack with 180 degrees of rotation (CK); Jeté with turn (JWT)) were evaluated. Male gymnasts had significant higher values in each anthropometric measure than females (p<0.01) except for sitting height/stature ratio, sitting height and fat mass (no significant differences). Female athletes were selected for their lowest fat mass. Height and ground contact time of technical leaps, squat jump and counter-movement jump, were significantly higher in male athletes than females. Height of hopping test (evaluating stiffness), and of difference between CMJ and squat jump (evaluating elastic properties of muscles) were no different between genders. Ground contact time of hopping test was significantly correlated (p<0.05) with ground contact time of technical leaps. Results of the present study suggest that similar anthropometric characteristics have to be researched for both gymnasts' gender. Reactivity and elastic muscle properties of the legs have to be research both in male and female athletes. [ABSTRACT FROM AUTHOR]

In this paper, we present a historical review of the whistle-smile reflex, a semiological sign missed in the literature and clinical practice., Competing Interests: The authors have no conflict of interests to declare., (Academia Brasileira de Neurologia. This is an open access article published by Thieme under the terms of the Creative Commons Attribution-NonDerivative-NonCommercial License, permitting copying and reproduction so long as the original work is given appropriate credit. Contents may not be used for commercial purposes, or adapted, remixed, transformed or built upon. (https://creativecommons.org/licenses/by-nc-nd/4.0/).)

Background: Hypericum perforatum (HP) is currently one of the most consumed medicinal plants in the world. In traditional Chinese medicine, the herb hypericum ( Guan Ye Lian Qiao ) belongs to the group of plants that clarify heat. It is also used to treat various types of infection and inflammation. In contrast to the extensive literature on the antidepressant effects of HP, little is known about its action on platelets. The main objective of this work was to investigate the possible relevance of HP to platelet function., Methods: We characterized the profile of platelet activation in the presence of HP extracts through an evaluation of molecular markers by flow cytometry: mobilization of intracellular Ca++ and expression of platelet receptors such as activated GPIIbIIIa and P-selectin (CD62)., Results: The results indicated a possible inhibitory effect of HP on the platelet activation response, which could be explained by the effect on intracellular calcium mobilization and the expression of activated GPIIbIIIa receptors. Despite of the limitations of an in vitro study, our results provide evidence of the possible mechanisms of action of HP., Conclusions: Further studies are needed to elucidate the effect of HP on hemostasis, but it may be recognized as a substance with antiplatelet properties.

Objective: Advance directives are becoming increasingly important as health technologies evolve. We sought to assess psychiatrists' knowledge of advance directives, as this knowledge is fundamental to the implementation and drafting of these personal documents., Methods: A previously published questionnaire that evaluated the knowledge of medical professors was used. The sample, composed of psychiatrists from Rio de Janeiro, Brazil, originated from a publicly available list. During the search process, the COVID-19 pandemic affected Brazil and the rest of the world, which influenced our methodology and results., Results: A total of 40 psychiatrists participated in the study. The results obtained, although not significant, suggested that psychiatrists with an increased time of practice had more knowledge of advance directives. Nevertheless, less than half of the participants had knowledge about this topic., Conclusion: The number of psychiatrists surveyed indicates the need for further studies on the subject. The influence of the COVID-19 pandemic on this study led to findings such as a change in sensitivity when addressing the topic and greater difficulty in contacting professionals., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Domingues, Castro, Monteiro, Cordero da Silva, Rego and Rego.)

Three patients with eosinophilic meningitis (EoM) were investigated in two hospitals in Porto Alegre, Southern Brazil. These patients had a common exposure after the ingestion of raw mollusks in a religious ritual. Two of them had an uncommon presentation with intense lower distal extremities pain and small fiber neuropathy as defined by an electroneuromyography (ENMG) study. All three patients were positive for Angiostrongylus cantonensis serology and recovered after antihelminthic and anti-inflammatory treatment. Increased awareness of A. cantonensis infection is important to avoid new infections and to improved recognition and handling of cerebral angiostrongyliasis., (Copyright © 2020 Elsevier B.V. All rights reserved.)

We describe a case of probable sporadic Creutzfeldt-Jakob disease in the setting of well-controlled HIV and discuss whether exist, in fact, HIV-related factors that may predispose to the development of prion disease. To the best of our knowledge, this is the third report of this association.

The aim of the study was to compare the psychophysical effects of rock climbing with a supervised fitness training in adults. Thirty-three healthy participants (M age=32 yr., SD=7) participated in rock climbing or in fitness training. The participants' functional fitness, anxiety, and mood states were tested before and after 3 mo. of training. There was significant improvement of physical fitness in both groups after the intervention period. Anxiety significantly decreased after each single training session at the end of both courses. Differential effects in the rock-climbing group, as compared to the fitness group, emerged only on Vigor. Specifically, the rock-climbing group showed a decreasing trend in Vigor while the fitness group showed an increasing trend of Vigor after the intervention.

Obstructive sleep apnoea syndrome (OSAS) is a sleep related breathing disorder caused by pharynx obstruction that often terminates in abrupt arousals and is capable of disrupting physiological sleep profile. Its' severity has been associated, among others, with craniofacial skeletal morphology. To investigate this relationship and elucidate craniofacial skeleton patterns in individuals without obvious maxillofacial abnormalities, 171 OSAS patients were studied with nocturnal polysomnographic record and cephalometric X-ray (24 variables). Cephalometric variables were compared between three apnoea/hypopnoea index (AHI) groups (AHI ≤ 15; 15 < AHI < 30; AHI ≥ 30) and uni/multivariate analysis between cephalometric variables and AHI were performed. The patients were predominantly men (83%), with a mean age of 48.1 years. Mean BMI and AHI were 28.4 kg/m(2) and 26.2, respectively. Most cephalometric variables differed among the three AHI groups. Fifteen cephalometric variables showed a correlation with AHI. Five cephalometric variables and BMI were independent AHI predictors. Cephalometric variables were better AHI predictors in normal weight patients. Significant evidence of craniofacial skeleton influence was found on OSAS severity, caudalization of the hyoid and lower sagittal facial projection being the most important patterns. From the cephalometric variables analysed, the hypopharynx calibre demonstrated a higher predictive value for AHI, independently of BMI., (Copyright © 2013 European Association for Cranio-Maxillo-Facial Surgery. Published by Elsevier Ltd. All rights reserved.)

Aim: Little is known about the physiological response during slideboard exercise (SE). The aim of the present study was to analyse the oxygen uptake (V.O2), the heart rate (HR) and the energy expenditure (EE) during a typical slideboard exercise session and investigate differences on these variables when performing the same choreography at two different cadences (130 e 145 beats per minute - bpm)., Methods: The sample comprised 13 female university students (21,77+/-0,97 years), apparently healthy and physically active, with past training in SE and mastering the technical levels 1 and 2. The subjects performed randomly exercise sessions at 130 bpm and 145 bpm. The ventilatory response was measured by an open air circuit system (COSMED K4b2, Rome, Italy) and HR was measured by a portable monitor (Polar Wireless Double Electrode, Kempele, Finland). HR and V.O2, during SE at 130 bpm, were 179.88+/-834 bpm and 37.95+/-3.71 mL/kg/min respectively. At 145 bpm SE mean values were 182.08+/-9.58 bpm and 39.67+/-3.82 mL/kg/min respectively. EE during 130 bpm exercise was 10.60+/-1.69 kcal/min and at 145 bpm was 10.90+/-1.36 kcal/min. No differences were found between 130 and 145 bpm in none of the variables. We conclude that slideboard exercise cardio respiratory response does not seem affected by the rhythm of execution. Moreover the EE associated with this type of exercise is above the literature reports for other types of group aerobic exercises.

Objective: To identify the incidence, risk factors, and mortality of acute renal failure (ARF) in patients undergoing myocardial coronary artery bypass surgery with extracorporeal circulation., Methods: All patients undergoing myocardial coronary artery bypass surgery were prospectively studied, and their clinical and laboratory variables were assessed using uni- and multivariate analysis (logistic regression)., Results: Acute renal failure occurred in 16.1% of the 223 patients studied; 4.9% of patients required dialysis. Risk factors associated with ARF in the univariate analysis were age > 63 years, OR 3.6 (95% CI=1.6 to 8.3); preoperative serum creatinine > 1.2 mg/dL, OR 5.9 (95% CI=2.4 to 14.6); duration of extracorporeal circulation > 90 min, OR 2.1 (95% CI=1.0 to 4.4); use of intraaortic balloon, OR 2.6 (95% CI=1.2 to 5.5); need for inotropic drugs, OR 4.4 (95% CI=1.9 to 10.2). In the multivariate analysis, independent factors associated with ARF were: age > 63 years, OR 3.0 (95% CI=1.3 to 7.2); preoperative serum creatinine > 1.2 mg/dL, OR 4.3 (95% CI=1.6 to 11.4); need for inotropic drugs, OR 3.2 (95% CI=1.3 to 8.0). Mortality in the patients with ARF was 25.0% compared with 1.1% in those without ARF and 63.6% in those who required dialysis., Conclusion: Acute renal failure after myocardial coronary artery bypass surgery is a frequent complication associated with a high mortality rate. The independent risk factors are age, previous renal failure, and the need for inotropic drugs.

Flow cytometry provides a convenient method to evaluate platelet activation by following the kinetics of intracellular free Ca2+, using sensitive fluorescent indicators that can be loaded into intact cells. Moreover, in the clinical setting, whole-blood techniques have obvious advantages to avoid artifactual platelet activation and allow the maintenance of near-physiological conditions. This unit describes a fast and sensitive flow cytometric procedure using the Ca2+-sensitive dye fluo-3 AM and the platelet-specific antibody CD41-PE to determine the kinetics of intracellular Ca2+ mobilization in whole-blood platelets with minimal manipulation of the samples. The technique may be applied to reveal fast and transient increases in cytosolic calcium upon platelet stimulation with the agonists ADP and thrombin. This protocol provides a simple and sensitive tool to assess in vitro the time course and intensity of signal-transduction responses to agonists under near-physiological conditions, and should be broadly applicable to studies of platelet reactivity.

Flow cytometry (FCM) allows the simultaneous measurement of multiple fluorescences and light scatter induced by illumination of single cells or microscopic particles in suspension, as they flow rapidly through a sensing area. In some systems, individual cells or particles may be sorted according to the properties exhibited. By using appropriate fluorescent markers, FCM is unique in that multiple structural and functional parameters can be quantified simultaneously on a single-particle basis, whereas up to thousands of biological particles per second may be examined. FCM is increasingly used for basic, clinical, biotechnological, and environmental studies of biochemical relevance. In this critical review, we summarize the main advantages and limitations of FCM for biochemical studies and discuss briefly the most relevant parameters and analytical strategies. Graphical examples of the biological information provided by multiparametric FCM are presented. Also, this review contains specific sections on flow cytoenzymology, FCM analysis of isolated subcellular organelles, and cell-free FCM.