A lack of efficacy of continuous theta burst stimulation over the left dorsolateral prefrontal cortex in autism: A double blind randomized sham-controlled trial.

Although previous open-label trials suggest the therapeutic potential of inhibitory repetitive transcranial magnetic stimulation (rTMS) over the dorsolateral prefrontal cortex (DLPFC) in autism spectrum disorder (ASD), methodological caveats exist. We conducted an 8-week randomized, double-blind sham-controlled trial to investigate the efficacy of inhibitory continuous theta burst stimulation (cTBS, a variant of rTMS) over the left DLPFC in individuals with ASD. Sixty children, adolescents and young adults (aged 8-30 years) with ASD without co-occurring intellectual disabilities were randomized to a 16-session 8-week cTBS versus sham stimulation course, with a follow-up 4 weeks after the trial. The Active group was not superior to the Sham group in any clinical or neuropsychological metrics at Week 8 or Week 12. Time effects of 8-week cTBS on symptoms and executive function were remarkable in both Active and Sham groups, with comparable response rates and effect sizes of changes in symptoms/cognition between groups. Our results from a sufficiently powered sample do not endorse the superior efficacy of cTBS over the left DLPFC to the shamed stimulation for children, adolescents and adults with ASD. These findings suggest that earlier positive open-label trial findings may be generalized by generalized/placebo effects. This highlights the urgent need for more rTMS/TBS studies with rigorous trial designs in ASD.

Risk modeling of single-cell transcriptomes reveals the heterogeneity of immune infiltration in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most common primary hepatic malignancies. E2F transcription factors play an important role in the tumorigenesis and progression of HCC, mainly through the RB/E2F pathway. Prognostic models for HCC based on gene signatures have been developed rapidly in recent years; however, their discriminating ability at the single-cell level remains elusive, which could reflect the underlying mechanisms driving the sample bifurcation. In this study, we constructed and validated a predictive model based on E2F expression, successfully stratifying patients with HCC into two groups with different survival risks. Then we used a single-cell dataset to test the discriminating ability of the predictive model on infiltrating T cells, demonstrating remarkable cellular heterogeneity as well as altered cell fates. We identified distinct cell subpopulations with diverse molecular characteristics. We also found that the distribution of cell subpopulations varied considerably across onset stages among patients, providing a fundamental basis for patient-oriented precision evaluation. Moreover, single-sample gene set enrichment analysis revealed that subsets of CD8+ T cells with significantly different cell adhesion levels could be associated with different patterns of tumor cell dissemination. Therefore, our findings linked the conventional prognostic gene signature to the immune microenvironment and cellular heterogeneity at the single-cell level, thus providing deeper insights into the understanding of HCC tumorigenesis.

The Effects of Virtual Reality Training on Balance, Gross Motor Function, and Daily Living Ability in Children With Cerebral Palsy: Systematic Review and Meta-analysis.

BACKGROUND: The increasing number of children with cerebral palsy (CP) has a serious impact on individuals, families, and society. As a new technology, virtual reality (VR) has been used in the rehabilitation of children with CP. OBJECTIVE: This study aimed to systematically evaluate the effect of VR training on balance, gross motor function, and daily living ability in children with CP. METHODS: PubMed, Embase, The Cochrane Library, Web of Science, and China National Knowledge Infrastructure databases were searched by computer, with the search period being from the establishment of each database to December 25, 2021, to collect randomized controlled trials (RCTs) on the effects of VR training on balance, gross motor function, and daily living ability in children with CP. The Cochrane risk of bias assessment tool was used to conduct quality assessment on the included literature, and RevMan software (version 5.3) was used to analyze data. RESULTS: A total of 16 articles were included, involving 513 children with CP. VR training can improve the balance function (Pediatric Balance Scale: mean difference 2.06, 95% CI 1.15-2.97; P<.001; Berg Balance Scale: mean difference 3.66, 95% CI 0.29-7.02; P=.03) and gross motor function (standardized mean difference [SMD] 0.60, 95% CI 0.34-0.87; P<.001) of children with CP. However, there is still certain disagreement on the impact on daily living ability (SMD 0.37, 95% CI -0.04 to 0.78; P=.08); after removing the source literature with heterogeneity, VR training can improve the daily living ability of children with CP (SMD 0.55, 95% CI 0.30-0.81; P<.001). CONCLUSIONS: VR training can significantly improve the balance function and gross motor function of children with CP, but the effect on the daily living ability of children with CP remains controversial.

Cascade Reaction of 2-Naphthols and Azirines: One-Pot Synthesis of C-3 Naphthol-Substituted Benzo[e]indoles.

A copper-catalyzed annulation of 3-aryl-2H-azirines with 2-naphthols has been developed for the rapid assembly of C-3-naphthol-substituted benzo[e]indoles in one pot. This cascade reaction was realized through dearomatic nucleophilic ring opening of azirine, intramolecular cyclization, and oxidative cross-dehydrogenative coupling to furnish the important unreported π-expanded naphthol/benzo[e]indole biaryls.

A Meta-Analysis: Intervention Effect of Mind-Body Exercise on Relieving Cancer-Related Fatigue in Breast Cancer Patients.

OBJECTIVE: This paper aims to systematically evaluate the intervention effect of mind-body exercise on cancer-related fatigue in breast cancer patients. METHODS: Databases including PubMed, the Cochrane Library, Embase, Web of Science, CNKI, Wanfang Data, and SINOMED were retrieved to collect randomized controlled trials on the effects of mind-body exercise on relieving cancer-related fatigue in breast cancer patients. The retrieval period started from the founding date of each database to January 6, 2021. Cochrane bias risk assessment tools were used to evaluate the methodological quality assessment of the included literature, and RevMan 5.3 software was used for meta-analyses. RESULTS: 17 pieces of researches in 16 papers were included with a total of 1133 patients. Compared with the control group, mind-body exercise can improve cancer-related fatigue in breast cancer patients. The combined effect size SMD = 0.59, 95% CI was [0.27, 0.92], p < 0.00001. Doing Tai Chi for over 40 minutes each time with an exercise cycle of ≤6 weeks can improve cancer-related fatigue in breast cancer patients more significantly. Sensitivity analysis shows that the combined effect results of the meta-analysis were relatively stable. CONCLUSION: Mind-body exercise can effectively improve cancer-related fatigue in breast cancer patients.

Biomimetic Ca2+ nanogenerator based on ions interference strategy for tumour-specific therapy.

Intracellular Ca2+ ions as second messenger played key role in cell behaviour, which was often overlooked in traditional antitumor treatment. Disrupting Ca2+ ion homeostasis by Ca2+ overload might switch ions signal from 'regulating' to 'destroying'. Inspired by this, a biomimetic Ca2+ nanogenerator was constructed. Briefly, the curcumin (CUR) was loaded into mesoporous calcium carbonate nanoparticles (MCC NPs), and then coated with platelet (PLT) membrane. Upon reaching tumour cells by PLT membrane-mediated tumour targeting effect, PLT@MCC/CUR would instantaneously decompose in acidic lysosomes, concurrently accompanying with Ca2+ generation and CUR release. The CUR could further facilitate Ca2+ release from endoplasmic reticulum (ER) and inhibit Ca2+ efflux, aggravating Ca2+ overload to disrupt mitochondrial Ca2+ homeostasis for mitochondria apoptosis signalling pathway activation. Interestingly, such effect was ineffective in normal cells, realising the tumour-specific therapeutic therapy. Based on ions interference strategy, PLT@MCC/CUR herein offered synergistic combination of Ca2+ overload therapy and chemotherapy, which would pave the way towards more effective nanotherapeutics.

Graft-derived extracellular vesicles transported across subcapsular sinus macrophages elicit B cell alloimmunity after transplantation.

Despite the role of donor-specific antibodies (DSAs) in recognizing major histocompatibility complex (MHC) antigens and mediating transplant rejection, how and where recipient B cells in lymphoid tissues encounter donor MHC antigens remains unclear. Contrary to the dogma, we demonstrated here that migration of donor leukocytes out of skin or heart allografts is not necessary for B or T cell allosensitization in mice. We found that mouse skin and cardiac allografts and human skin grafts release cell-free donor MHC antigens via extracellular vesicles (EVs) that are captured by subcapsular sinus (SCS) macrophages in lymph nodes or analog macrophages in the spleen. Donor EVs were transported across the SCS macrophages, and donor MHC molecules on the EVs were recognized by alloreactive B cells. This triggered B cell activation and DSA production, which were both prevented by SCS macrophage depletion. These results reveal an unexpected role for graft-derived EVs and open venues to interfere with EV biogenesis, trafficking, or function to restrain priming or reactivation of alloreactive B cells.

Predictors and one-year outcomes of patients with delayed graft function after deceased donor kidney transplantation.

BACKGROUND: Delayed graft function (DGF) is closely associated with the use of marginal donated kidneys due to deficits during transplantation and in recipients. We aimed to predict the incidence of DGF and evaluate its effect on graft survival. METHODS: This retrospective study on kidney transplantation was conducted from January 1, 2018, to December 31, 2019, at the Second Xiangya Hospital of Central South University. We classified recipients whose operations were performed in different years into training and validation cohorts and used data from the training cohort to analyze predictors of DGF. A nomogram was then constructed to predict the likelihood of DGF based on these predictors. RESULTS: The incidence rate of DGF was 16.92%. Binary logistic regression analysis showed correlations between the incidence of DGF and cold ischemic time (CIT), warm ischemic time (WIT), terminal serum creatine (Scr) concentration, duration of pretransplant dialysis, primary cause of donor death, and usage of LifePort. The internal accuracy of the nomogram was 83.12%. One-year graft survival rates were 93.59 and 99.74%, respectively, for the groups with and without DGF (P < 0.05). CONCLUSION: The nomogram established in this study showed good accuracy in predicting DGF after deceased donor kidney transplantation; additionally, DGF decreased one-year graft survival.

Effects of non-caloric ultrashort wave on the expression of CoQ10 and C1GALT1C1 in rats with cerebral ischemia reperfusion injury. / æ— çƒ­é‡è¶ çŸ­æ³¢å¯¹è„‘ç¼ºè¡€å†çŒæ³¨æŸä¼¤å¤§é¼ æ—©æœŸCoQ10和C1GALT1C1表达的影响.

OBJECTIVES: To examine the changes of coenzyme Q10 (CoQ10) and ß1,3-galactosyl transferase specific chaperone 1 (C1GALT1C1) in brain of rats with ischemic injury at different time points and to explore the protective mechanism of ultrashort wave (USW) on ischemic brain injury. METHODS: Fifty SD rats were randomly divided into 5 groups (n=10 per group): a sham group (control group) and 4 experimental group (ischemia for 2 h). The 4 experimental groups were set as a model 1 d group, a USW 1 d group, a model 3 d group and a USW 3 d group, respectively. Five rats were randomly selected for 2,3,5-triphenyltetrazoliumchloride (TTC) staining in each experimental group, and the remaining 5 rats were subjected to Western blotting and real-time PCR. The percentage of cerebral infarction volume and the relative expression level of CoQ10 and C1GALT1C1 in the brain were examined and compared. RESULTS: The infarct volume percentage after TTC staining was zero in the sham group. With the progress of disease and USW therapy, the infarct volume percentage was decreased in the experimental groups (all P<0.05); Western blotting and real-time PCR showed that the relative expression level of CoQ10 in the sham group was the highest, while in the experimental groups, the content of CoQ10 showed a upward trend with the extension of disease and USW therapy, with significant difference (all P<0.05). The relative expression level of C1GALT1C1 in the sham group was the lowest, but in the experimental groups, they showed a downward trend with the extension of disease and USW therapy, with significant difference (all P<0.05). CONCLUSIONS: Non-caloric USW therapy may upregulate the expression of CoQ10 to suppress the expression of C1GALT1C1 in rats, leading to alleviating cerebral ischemic reperfusion injury.

[Multilevel Modelling on the Prevalence and Determinants of Depressive Symptoms in Middle and Old-aged Rural-to-urban Immigrants in Chengdu].

OBJECTIVE: To determine the prevalence and determinants of depressive symptoms in middle and old-aged rural-to-urban immigrants in Chengdu. METHODS: A total of 1 645 middle and old-aged rural-to-urban immigrants aged over 45 yr. were selected to participate in a questionnaire survey through a multi-stage random sampling method in Chengdu. Multilevel (households and individuals) models were established to identify predictors of depressive symptoms. RESULTS: About 14.5% of respondents reported depressive symptoms. The multilevel model indicated that family clustering of depressive symptoms existed. Household income and length of urban life at the household level, and age, chronic diseases, smoking, and social support at the individual level were significant predictors of depressive symptoms. CONCLUSION: The prevalence of depressive symptoms in middle and old-aged rural-to-urban immigrants deserves increasing policy attention for the purpose of promoting mental health in the population.

Robot-assisted laparoscopic vs laparoscopic donor nephrectomy in renal transplantation: A meta-analysis.

BACKGROUND: Currently, the robot-assisted laparoscopic donor nephrectomy (RDN) technique is used for live donor nephrectomy. Does it provide sufficient safety and benefits for living donors? We conducted a meta-analysis to assess the safety and efficacy of RDN compared with the laparoscopic donor nephrectomy (LDN). MATERIAL AND METHODS: Eligible studies were retrieved and screened from electronic databases from 1999 onward: PubMed, Cochrane Library, and Web of Science. Relevant parameters were explored using Review Manager V5.3 and included operative time, warm ischemia time, estimated blood loss, and length of hospital stay. RESULTS: Compared with RDN, LDN had shorter operative time (min; weighted mean difference (WMD): -0.53; 95% CI: [-0.85, 0.20]; P = 0.001) and warm ischemia time (second; WMD: -55.01; 95% CI: [-71.56, 38.45]; P < 0.00001) and less estimated blood loss (mL; WMD: -28.30, 95% CI: [-46.37, 10.24], P = 0.002). The pooled analysis of postoperative pain showed lower visual analog scale (VAS) scores for RDN compared with LDN (WMD:1.28, P < 0.00001). We also observed that length of hospital stay, postoperative serum creatinine (SCr) in donors, postoperative estimated glomerular filtration rate (eGFR) of recipients and postoperative complications for donors were not significantly different between groups. CONCLUSION: As long as RDN is practiced proficiently, it is believed that RDN is a feasible alternative to LDN.

Study on the equity of medical services utilization for elderly enrolled in different basic social medical insurance systems in an underdeveloped city of Southwest China.

BACKGROUND: The equity of medical services utilization for elderly individuals enrolled in different basic social medical insurance systems holds significant meaning for social harmony against a background of demographic aging and a growing wealth gap in China. This study is to explore the equity of the three medical insurance systems in southwest China with the aim of providing recommendations for relevant policy. METHODS: A total of 9600 elderly people insured through basic social medical insurance were selected and interviewed with a questionnaire. This study used a binary logistic regression model to investigate the effect of household income for medical services utilization and adopted a concentration index to measure the inequity of medical services utilization among elderly participants enrolled in different medical insurance categories. RESULTS: Outpatient services utilization was almost identical in the different insurance systems (78.5%, 77.7% and 78.6%). There were no statistically significant differences according to income level in the Urban Employee Basic Medical Insurance (UEBMI) and Urban Resident Basic Medical Insurance (URBMI) programs, but in the New Cooperative Medical Scheme (NCMS), higher-income groups tended to utilize more services. The corresponding concentration index (CI) values were 0.0162, 0.0173 and 0.0179 respectively. The NCMS showed a lower hospitalization rate than UEBMI and URBMI (17.7% vs 24.2% and 24.9%). The higher income group utilized hospitalization more, regardless of the insurance system. The corresponding CI values were 0.0817, 0.0605 and 0.0319 respectively. CONCLUSION: The equity of medical services utilization for elderly exist in all three health insurance systems, in particular, the inequities in utilization of hospitalization were more severe than outpatient services. UEBMI and URBMI were better than NCMS in the equity of outpatient services. Although NCMS was more equitable than URBMI and UEBMI in terms of hospitalization, this was based on "overall low utilization of hospitalization regardless of income levels" in NCMS compared with URBMI and UEBMI. The disparities of the three basic social medical insurance systems should be eliminated. For low-income residents, specific insurance policies including reducing deductible, covering more medical service and increasing reimbursement ratio could be considered.

[Medical Service Information Seeking Behaviors in Rural and Urban Patients in Sichuan Province].

OBJECTIVE: To understand how rural and urban patients seek medical service information in Sichuan province. METHODS: A self-designed questionnaire was distributed randomly to patients who visited primary,secondary and tertiary health facilities in Chengdu,Yibin and Suining,collecting data in relation to their sources of medical service information,as well as the contents and credibility of the information. RESULTS: The major sources of medical service information came from friends,past experiences and television programs,which were consistent with the most desirable access channels. The urban patients were more likely to trust (5.3%) and use (10.6%) the Internet to obtain medical service information compared with their rural counterparts (3.4% and 5.5%,respectively,P<0.05). The most sought after information concerned about medical staff,reputation and price. The rural patients were more likely to be concerned about price (26.7%) than their urban counterparts (20.3%,P<0.05). The choices of patients were likely to be influenced by advices from family members,relatives and friends,and doctors and nurses. The patients had a higher level of trust in doctors and nurses than their relatives and friends,but lower than their family members. CONCLUSION: Patient choices are shaped by their medical service information seeking behaviors and advices from others. Targeted marketing strategies for urban and rural patients should be developed to channel patients to appropriate health facilities.

[Health Determinants of Rural Elderly Populations with Different Living Arrangements: a Multilevel Model].

OBJECTIVE: To determine the health and its determinants of rural elderly populations in a city in Sichuan province. METHODS: Data were extracted from a 2015 household survey, containing 7 280 rural participants aged over 60 years. The prevalence of two-week morbidity and chronic conditions were compared among those with different living arrangements. Multilevel models were established to determine the family and individual factors associated with the health status of the respondents. RESULTS: Most respondents (57.0%) lived with a spouse, compared with 15.4% who lived alone. Those who lived alone had higher prevalence of two-week morbidity (48.0%) and chronic conditions (70.0%). The multilevel model indicated a familial clustering effect. Living arrangements and household income were associated with the prevalence of two-week morbidity and chronic conditions, after controlling for variations in age, depression, smoking, drinking, and exercise at the individual level. CONCLUSIONS: The rural elderly people who live alone have poor health status. Household income, age, depression, smoking, drinking and physical exercise are associated with two-week morbidity and chronic conditions.

[Overweight and Obesity among New Urban Migrants in Chengdu and Kunming].

OBJECTIVE: To determine the prevalence of overweight and obesity in new urban migrants. METHODS: A questionnaire survey was conducted on 21 651 urban residents in the downtown and surrounding urban areas of Kunming and Chengdu selected through a multi-stage stratified cluster sampling method. Factors associated with overweight and obesity were identified using Chi-square tests and multivariate logistic regression models. RESULTS: 11 626 people were enrolled. The new urban migrants (n=7 555) had 31.2% overweight and 6.7% obesity, compared with 24.9% and 4.3%, respectively, in the urban residents(n=4 071). Gender, age, educational, marital status and alcohol consumption were associated with overweight and obesity in the new urban migrants. Gender, age, education and alcohol consumption were predictors of overweight and obesity of the urban residents. CONCLUSIONS: Urban new immigrants are more likely to be overweight and obese than urban residents, although predictors for overweight and obesity remain similar.

Establishment of Chinese Cardiovascular Surgery Registry Database / 中国循环杂志

Objective: To introduce the construction idea and function for establishing China Cardiovascular Surgery Registry (CCSR)database and to provide a reference for domestic congener databases. Methods: Using peer database as reference, taking current status of cardiovascular surgery registry and hardware in our country with the necessity of international communication, we worked on a variables selection, metadata instruction, logic rules, case report form develpment and finally established a web-based, multi-functional database that enabled cross-database and international merging of data, forming a national intelligent data-exchanging platform for cardiovascular surgery. Results:CCSR database has over 300 variables of multiple topics including basic information, risk factors, medical procedures and endpoint events. Taking clinical and association data exchange standards as reference, it may conduct cross-discipline data connection, record important peri-operative information in relevant patients and meanwhile, it has the functions of automatic logic check, data report, statistical study, data export and importing the electronic medical records. Conclusion:CCSR database is a national platform accord with current status of Chinese cardiovascular surgery and characteristics, meanwhile it gives consideration to international communication and data exchange; which may play a important role in improving medical care and clinical investigation.

Establishment of Chinese Cardiovascular Surgery Registry Database / 中国循环杂志

Objective: To introduce the construction idea and function for establishing China Cardiovascular Surgery Registry (CCSR)database and to provide a reference for domestic congener databases. Methods: Using peer database as reference, taking current status of cardiovascular surgery registry and hardware in our country with the necessity of international communication, we worked on a variables selection, metadata instruction, logic rules, case report form develpment and finally established a web-based, multi-functional database that enabled cross-database and international merging of data, forming a national intelligent data-exchanging platform for cardiovascular surgery. Results:CCSR database has over 300 variables of multiple topics including basic information, risk factors, medical procedures and endpoint events. Taking clinical and association data exchange standards as reference, it may conduct cross-discipline data connection, record important peri-operative information in relevant patients and meanwhile, it has the functions of automatic logic check, data report, statistical study, data export and importing the electronic medical records. Conclusion:CCSR database is a national platform accord with current status of Chinese cardiovascular surgery and characteristics, meanwhile it gives consideration to international communication and data exchange; which may play a important role in improving medical care and clinical investigation.

[Research on chemotherapy measures in different Clonorchis sinensis infectiosity endemic areas].

OBJECTIVE: To explore the suitable chemotherapy measures in different Clonorchis sinensis infectiosity endemic areas. METHODS: The interventional groups and control groups were set up in the heavy, moderate and light C. sinensis infectiosity areas in Guangdong Province, respectively. In the intervention groups, the chemotherapy was administered among all the residents aged above 3 years in the heavy endemic areas, the chemotherapy was given among the focus populations in the moderate endemic areas, and the chemotherapy was administered among the residents infected with C. sinensis confirmed by stool examinations in the light endemic areas. No measures were carried out in the control groups. RESULTS: One year after the interventions, the C. sinensis infection rates of the interventional groups decreased by 47.90%, 86.52% and 100%, the abnormal rates of liver B ultrasonic examinations decreased by 26.50%, 31.00% and 100%, the ALT abnormal rates decreased by 48.70%, 62.30% and 100%, and the AST abnormal rates decreased by 55.50%, 39.90% and 100% in the heavy, moderate and light endemic areas, respectively. There was no decreasing trend of above-mentioned indexes in the control groups. CONCLUSION: In the different C. sinensis infectiosity endemic areas, the different chemotherapy measures should be carried out.

[Effect of health education intervention in schools of Yanrui Town, Yushan County].

OBJECTIVE: To evaluate the effect of a new mode of health education in schools. METHODS: In the Zhaiqian Primary School, Yanrui Town, Yushan County in a hilly schistosomiasis endemic area, a new mode of health education intervention, i. e. "rewards and punishment + advise others by using one's experience + teachers' participation" was carried out, and the knowledge, attitude and practice of schistosomiasis prevention of the pupils, and the schistosome infection rates of the pupils were investigated and the results were analyzed and compared before and after the intervention. RESULTS: Among 204 pupils investigated, the awareness rate of schistosomiasis prevention knowledge increased from 26.47% before the intervention to 86.76% and 99.51% one and two years after the intervention, respectively; the rate of correct attitude increased from 17.04% to 73.04% and 100%, respectively; the rate of the infested water contact decreased from 83.33% to 26.96% and 0, respectively; the schistosome infection rate decreased from 2.94% to 0.49% and 0, respectively. CONCLUSION: The health education of schistosomiasis prevention in schools is necessary, and the new mode of health education, "rewards and punishment + advise others by using one's experience + teachers' participation" is effective in the prevention from schistosome infection in pupils.

LC-MS/MS determination of helicid in human plasma and its application in pharmacokinetic studies.

Helicid is a traditional Chinese medicine used to treat headache and insomnia with definite effects. To facilitate pharmacokinetic studies of helicid in man, a sensitive and specific LC-MS/MS method for the quantitative detection of helicid in human plasma was developed and validated. The method involved the addition of bergeninum as the internal standard (IS), protein precipitation, HPLC separation, and quantification by MS/MS system using negative electrospray ionization in the multiple reaction monitoring mode (MRM). The precursor→product ion transitions were monitored at m/z 282.8→120.9 for helicid and m/z 326.9→192.2 for the IS, respectively. The lower limit of quantification (LLOQ) was 0.2 µg/L. The calibration curves for helicid was linear over a concentration range of 0.2-20 µg/L. The intra- and inter-batch analyses of QC samples at 0.4, 2, 20 µg/L indicated good precision (%R.S.D. between 2.69 and 5.47%) and accuracy (between 96.15 and 105.05%). The helicid was stable in human plasma stored at room temperature for at least 24h, 4°C for at least 24h, -20°C for at least 1 month, and for routine three freeze-thaw cycles. This accurate and specific assay provides a useful method for evaluating the pharmacokinetic profile of helicid in humans.