An optimized protocol for detecting guard cell specific gene expression by in situ RT-PCR in Brassica rapa

It is important to detect specific genes expressed in the guard cells,which control gas exchange and play key roles in response to drought and salt stresses.Due to the genetic transformation of Chinese cabbage(Brassica rapa)has not been well developed,in situ RT-PCR is a valuable option for detecting guard cell specific genes.We reported an optimized protocol of in situ RT-PCR by using an FAMA homologous gene Bra001929 in Brassica rapa.FAMA in Arabidopsis has been verified to be specially expressed in guard cells.We designed specific RT-PCR primers and optimized the protocol in terms of the(a)reverse transcription time,(b)blocking time,(c)antigen-antibody incubation time,and(d)washing temperature.Our approach provides a sensitive and effective in situ RT-PCR method for locating expression in the guard cells in Brassica rapa.Moreover,we proved the guard cell specific expression of Bra001929 in the epidermis indicating its’applicability as a marker gene for guard cells of Brassica rapa.

A STUDY ON EXPRESSION OF ARmRNA IN HUMEN BPH TISSUE WITH IN SITU RT-PCR

Objecive To check and compare the expression levels of human androgen receptor (AR) mRNA in both normal adult prostate (NAP) and benign prostate hyperplasia (BPH) tissues, and to try to find if BPH results from the abnormal transcription of ARmRNA in prostate. Methods Expression of the human ARmRNA in 14 paraffin-embedded prostate tissues (4 cases of NAP and 10 cases of BPH) was studied with the direct in situ reverse transcription-polymerase chain reaction (RT-PCR). Quantitative analysis of the ARmRNA products was performed using the image analysis system. Results ①Specific ARmRNA was detected in both NAP and BPH specimens and in both epithelia and interstitial cells. The positive products were relatively densely localized in the cytoplasm of perinuclear zone. ② The intensity of ARmRNA signals in epithelial cells was significantly stronger than that in interstitial cells (P<0.001). However, there was no statistically significant difference in ARmRNA level between NAP and BPH; ③ The heterogeneity of ARmRNA signal and the androgen-independent cells were observed in prostatic epithelia. Stronger positive signals of ARmRNA were shown in a few basal-cell layer (BCL) cells of BPH tissue, but were not found in that of NAP tissue. Conclusion Results of this study show that there is no significant difference in the ARmRNA expression between NAP and BPH groups in both epithelium and interstitial cells. It may indicate that BPH does not result from the ARmRNA transcription in the prostate.

A Study of the Distribution of Apple stem pitting virus in Tissues of Pear Tree Using In Situ Hybridization and In Situ RT-PCR

To understand the distribution of Apple stem pitting virus （ASPV） in tissues of pear tree and provide directly theorical and technical support for shoot tips detoxication, leaves and shoot tips of Korla pear were used as the materials, cDNA probe for ASPV was synthesized through RT-PCR reaction system using the unradioactive digoxigenin-labeled probe, and the specificity and sensitivity of probe were verified by blot hybridization method. Paraffin slice for in situ PCR and in situ hybridization was made and the location and distribution of ASPV RNA were detected in paraffin slices using in situ reverse transcription polymerase chain reaction, and the important factors which influenced the experiment results were optimized. ASPV mainly distributed in palisade tissue of mesophyll cells, external cortex of the tip, and the corresponding newborn vascular bundles. 20 min was the suitable digestive time for proteinase K. For the better amplication, RT reaction system should be above 0.2 U μL^-1 and 0.4 mmol L^-1 for RNasin and dNTPs respectively, 0.1-1.3 U μ L^-1 SuperScript Ⅱ, 0.6- 0.8 μmol L^-1 primer concentration, and above 0.5 U 100 μL^-1 LA Taq DNA polymerase. The suitable annealing temperature in PCR reaction was 60℃ with 35 cycles. The apical meristem of 0.25 mm was the region of virus-free.

蓝舌病新疆分离株VP7蛋白编码基因序列分析及一步法RT-PCR方法的建立

【目的】分析蓝舌病新疆分离株编码VP7蛋白S7基因保守序列,建立蓝舌病新疆分离株一步法RT-PCR方法,为新疆BTV分子流行病学调查及防控提供技术支持。【方法】采用二代测序技术获得新疆分离株S7基因序列,登陆GenBank对序列进行同源性Blast比对,用软件MEGA 5.0分析序列差异,根据蓝舌病新疆分离株编码VP7蛋白S7基因保守序列,运用软件Oligo6.0设计引物,对蓝舌病新疆分离株S7基因进行RT-PCR扩增,并验证该方法的特异性及敏感性。【结果】蓝舌病中国新疆分离株编码VP7蛋白S7基因序列与哈尔滨报道BTV分离株S7基因片段相似度较高为89.64%,与中国云南报道BTV-29型分离株、蒙古国BTV分离株S7基因片段相似度分别为87.49%和87.39%;与德国BTV分离株S7基因片段相似度为80.70%,其余均无同源性序列。中国新疆分离株S7基因片段序列与4条同源序列间存在26处核苷酸差异,9处氨基酸差异。【结论】建立的蓝舌病中国新疆分离株S7基因片段一步法RT-PCR方法具有良好的特异性,仅BTV中国新疆分离株扩增获得目的条带,该方法的敏感性为4.0×10^(3)copies/mL。

猪Linda病毒巢式RT-PCR检测方法的建立

猪Linda病毒能引起仔猪先天性震颤,严重危害养猪业的发展,目前在我国暂未发现感染病例。本研究根据猪Linda病毒Core-E~(ms)基因序列,设计并合成巢式RT-PCR引物,通过对退火温度、引物浓度等进行优化,建立了猪Linda病毒巢式RT-PCR检测方法。结果显示,该方法具有良好的特异性,与非洲猪瘟病毒、猪瘟病毒、猪繁殖与呼吸综合征病毒、猪伪狂犬病病毒、猪圆环病毒2型均无交叉反应;敏感性好,对慢病毒阳性对照品的最低检出限为10~1 copies/μL,比普通RT-PCR灵敏10倍。使用该方法检测模拟病毒样品,发现最低检出限为10~1 TU/mL,与荧光定量RT-PCR方法一致。本研究首次建立了可检测猪Linda病毒的巢式RT-PCR方法,其特异性强、灵敏度高,为在口岸以及条件有限的场地对猪Linda病毒进行精准且快速的检测提供了有效技术手段,也为防范Linda病毒传入提供了有力技术支撑。

帕利亚姆病毒实时荧光定量RT-PCR检测方法的建立与应用

本研究拟建立帕利亚姆病毒(Palyam virus,PALV)血清型特异性实时荧光定量RT-PCR(qRT-PCR)方法用于临床样本或媒介中PALV血清型鉴定。根据我国流行PALV毒株的基因节段2序列,设计扩增引物和TaqMan探针,建立PALV血清型特异型qRT-PCR方法,对方法的特异性、灵敏性与重复性进行评估;以我国分离的28株PALV和90份核酸阳性血液样本评估检测方法的可靠性;利用建立的方法对采集库蠓样本中携带的PALV进行血清型鉴定。结果显示,建立的PALV血清型qRT-PCR检测方法具有良好的特异性与灵敏性,可检出核酸拷贝数下限在22至28 copies·μL^(-1)。对28株PALV的qRT-PCR检测结果与病毒测序鉴定结果一致;对PALV不同感染阶段哨兵动物血液(90份)中的qRT-PCR鉴定结果与分离病毒的血清型鉴定结果一致;建立的方法可准确鉴定库蠓中携带PALV的血清型。本研究建立的PALV血清型qRT-PCR定型方法具有良好的特异强、敏感性与重复性,可用于PALV感染动物与媒介中PALV血清型的鉴定,具有良好的应用价值。

基于RT-PCR方法对河北省桃病毒和类病毒种类的调查鉴定

病毒是造成桃果实产量和品质下降的原因之一,RT-PCR是检测病毒的常用分子生物学方法。利用RT-PCR方法对河北省113份桃样品进行了13种病毒的检测,结果表明,检测到的10种病毒包括ACLSV、PBNSPaV、HSVd、NSPaV、PNRSV、PaLV、PLMVD、APCLSV、PeVD和PPV,其中ACLSV检测率最高,其次是PBNSPaV和HSVd 2种病毒。病毒具有复合侵染的现象,三重侵染和四重侵染的检测率最高,没有检测到不被病毒侵染和单一侵染的样品。研究结果为河北省桃病毒脱除、无病毒苗木繁育奠定了理论基础。

鸽微RNA病毒实时荧光定量RT-PCR检测方法的建立

旨在建立鸽微RNA病毒(pigeon megrivirus,PiMeV)实时荧光定量RT-PCR检测方法。本研究根据GenBank中PiMeVs序列特征设计特异性检测引物,从信鸽粪便中检测到PiMeV阳性(命名为PiMeV-CHN001株),并对其3 C基因进行核苷酸同源性比较和遗传进化分析,明确其基因特征后,设计特异性实时荧光定量RT-PCR检测(RT-qPCR)引物组,建立检测PiMeV的RT-qPCR方法。结果显示:PiMeV-CHN001株3 C基因全长为591 bp,编码197个氨基酸,和其他2株野鸽源PiMeV(MeV-B1株和MeV-B2株)核苷酸相似性分别为89.5%和92.0%。建立的检测PiMeV的RT-qPCR方法的标准曲线Y轴截距为37.93,斜率为-3.335,相关系数为1.00,扩增效率为99.4%。特异性强,仅PiMeV出现特异性扩增信号和特异性峰值[Tm值为(81.69±0.22)℃],对鸽源禽流感病毒(avian influenza virus,AIV)、鸽源禽I型副黏病毒(pigeon paramyxovirus type I,PPMV-1)、鸽输血传播病毒(pigeon torque teno virus,PTTV)、鸽腺病毒(pigeon adenovirus,PiAd)及鸽圆环病毒(pigeon circovirus,PiCV)检测均未见特异性扩增信号;敏感性优,最低检测限为54.0拷贝·μL^(-1);重复性好,批内和批间变异系数均低于1.5%。用建立的检测方法对42份信鸽粪便样品进行检测,发现2份阳性样品(阳性率为4.76%)。本研究首次证实我国大陆地区信鸽中存在PiMeV,丰富了PiMeV宿主谱信息;建立的RT-qPCR方法为后续开展PiMeV流行病学研究提供支撑。

柑橘3种病毒类病原多重RT-PCR检测技术的建立及应用

【目的】建立柑橘黄化脉明病毒(citrus yellow vein clearing virus,CYVCV)、柑橘衰退病毒(citrus tristeza virus,CTV)和啤酒花矮化类病毒(hop stunt viroid,HSVd)的多重RT-PCR检测体系。【方法】设计多重RT-PCR引物,分析其特异性,确定其最佳浓度比、最适退火温度及灵敏度,在此基础上对福建地区的柑橘样品进行检测。【结果】确定了CYVCV-F/R、CTV-F/R和HSVd-F/R等3对引物的最佳浓度比例为1∶1∶2,最适退火温度为52.9℃,灵敏度结果显示该体系可检测模板稀释到10^(-2)的阳性样品。应用该体系对采自福建部分地区的157份柑橘样品进行检测,结果发现,CYVCV、CTV和HSVd的检出率分别为47.1%、56.7%和22.9%。【结论】成功建立了柑橘CYVCV、CTV和HSVd病原的多重RT-PCR检测方法,为该类病害的检测提供准确、快速的检测方法。

山羊地方性鼻内肿瘤病毒TaqMan荧光定量RT-PCR检测方法的建立及应用

山羊地方性鼻内肿瘤(enzootic nasal tumor, ENT)是山羊的病毒性传染病,由山羊地方性鼻内肿瘤病毒(enzootic nasal tumor virus of goats, ENTV-2)所引起,感染后可以导致山羊的鼻道筛骨上皮细胞发生不可逆的癌变,已在世界范围内广泛存在,对山羊养殖业造成严重的经济损失。为建立快速、准确且能定量分析ENTV-2的检测方法,本研究根据GenBank上发布的ENTV-2 env基因(MT254061.1)保守区域设计引物和TaqMan探针,建立ENTV-2 TaqMan探针荧光定量RT-PCR检测方法,并对其特异性、灵敏性、重复性与临床检测效果进行验证。结果显示,重组质粒标准品模板浓度为6.30×10^(2)~6.30×10^(7) copies·μL^(-1)呈现良好的线性关系,相关系数R^(2)为0.996 5,扩增效率为110%,线性方程的斜率为-2.953,最低检测限度为6.30×10^(1) copies·μL^(-1);组内变异系数和组间变异系数均小于2%,重复性好;与口蹄疫病毒((foot-and-mouth disease virus, FMDV)、小反刍兽疫病毒(peste des petits ruminants virus, PPRV)、蓝舌病毒(bluetongue virus)、羊内源性逆转录病毒(endogenous retroviruses)等均无交叉反应,表明该方法具有很好的特异性。利用本研究所建立的TaqMan探针实时荧光定量RT-PCR检测方法对29份临床样品进行检测,该方法较普通PCR方法的检出率更高。综上表明,本研究成功建立了ENTV-2 TaqMan探针实时荧光定量RT-PCR检测方法,为ENTV-2的快速诊断和流行病学调查提供技术手段。

Genesis of the Nuri Cu-W-Mo Deposit,Tibet,China:Constraints from in situ Trace Elements and Sr Isotopic Analysis of Scheelite

The Nuri deposit is the only Cu-W-Mo polymetallic deposit with large-scale WO3 resources in the eastern section of the Gangdese metallogenic belt,Tibet,China.However,the genetic type of this deposit has been controversial since its discovery.Based on a study of the geological characteristics of the deposit,this study presents mineralization stages,focusing on the oxide stage and the quartz-sulfide stage where scheelite is mainly formed,referred to as Sch-A and Sch-B,respectively.Through LA-ICP-MS trace element and Sr isotope analyses,the origin,evolutionary process of the oreforming fluid and genesis of the ore deposit are investigated.Scanning Electron Microscope-Cathodoluminescence(SEMCL)observations reveal that Sch-A consists of three generations,with dark gray homogenous Sch-A1 being replaced by relatively lighter and homogeneous Sch-A2 and Sch-A3,with Sch-A2 displaying a gray CL image color with vague and uneven growth bands and Sch-A3 has a light gray CL image color with hardly any growth band.In contrast,Sch-B exhibits a‘core-rim’structure,with the core part(Sch-B1)being dark gray and displaying a uniform growth band,while the rim part(Sch-B2)is light gray and homogeneous.The normalized distribution pattern of rare earth elements in scheelite and Sr isotope data suggest that the early ore-forming fluid in the Nuri deposit originated from granodiorite porphyry and,later on,some country rock material was mixed in,due to strong water-rock interaction.Combining the O-H isotope data further indicates that the ore-forming fluid in the Nuri deposit originated from magmatic-hydrothermal sources,with contributions from metamorphic water caused by water-rock interaction during the mineralization process,as well as later meteoric water.The intense water-rock interaction likely played a crucial role in the precipitation of scheelite,leading to varying Eu anomalies in different generations of scheelite from the oxide stage to the quartz-sulfide stage,while also causing a gradual decrease in oxygen fugacity(fO2)and a slow rise in pH value.Additionally,the high Mo and low Sr contents in the scheelite are consistent with typical characteristics of magmatic-hydrothermal scheelite.Therefore,considering the geological features of the deposit,the geochemical characteristics of scheelite and the O-H isotope data published previously,it can be concluded that the genesis of the Nuri deposit belongs to porphyry-skarn deposit.

高致病性猪蓝耳病RT-PCR检测

为了解当地生猪养殖场猪蓝耳病感染情况,这次调查研究和结果为高致病性猪蓝耳病的预防和控制提供了实验数据的依据。本实验就当地一些规模养猪场和一些个体养猪户猪场养的猪突然出现以食欲废绝、耳末梢发紫、体温高、发病率、死亡率高为特征的疾病,对病死猪进行了无菌采样,实验室检测。

Analytical investigations of in situ stress inversion from borehole breakout geometries

This study aims to investigate the feasibility of deriving in situ horizontal stresses from the breakout width and depth using the analytical method.Twenty-three breakout data with different borehole sizes were collected and three failure criteria were studied.Based on the Kirsch equations,relatively accurate major horizontal stress(sH)estimations from known minor horizontal stress(sh)were achieved with percentage errors ranging from 0.33%to 44.08%using the breakout width.The Mogi-Coulomb failure criterion(average error:13.1%)outperformed modified Wiebols-Cook(average error:19.09%)and modified Lade(average error:18.09%)failure criteria.However,none of the tested constitutive models could yield reasonable sh predictions from known sH using the same approach due to the analytical expression of the redistributed stress and the nature of the constitutive models.In consideration of this issue,the horizontal stress ratio(sH/sh)is suggested as an alternative input,which could estimate both sH and sh with the same level of accuracy.Moreover,the estimation accuracies for both large-scale and laboratory-scale breakouts are comparable,suggesting the applicability of this approach across different breakout sizes.For breakout depth,conformal mapping and complex variable method were used to calculate the stress concentration around the breakout tip,allowing the expression of redistributed stresses using binomials composed of sH and sh.Nevertheless,analysis of the breakout depth stabilisation mechanism indicates that additional parameters are required to utilise normalised breakout depth for stress estimation compared to breakout width.These parameters are challenging to obtain,especially under field conditions,meaning utilising normalised breakout depth analytically in practical applications faces significant challenges and remains infeasible at this stage.Nonetheless,the normalised breakout depth should still be considered a critical input for any empirical and statistical stress estimation method given its significant correlation with horizontal stresses.The outcome of this paper is expected to contribute valuable insights into the breakout stabilisation mechanisms and estimation of in situ stress magnitudes based on borehole breakout geometries.

一步RT-PCR法检测苹果锈果类病毒和苹果凹果类病毒

由类病毒引起的苹果病害在中国苹果主产区普遍发生,严重制约苹果产业的健康发展。对采自不同地区的带有典型症状的弘前富士、南方脆及惠民短枝富士果实,进行苹果锈果类病毒(ASSVd)和凹果类病毒(ADFVd)检测,发现两种病毒能够共侵染。根据ASSVd和ADFVd保守序列设计通用引物,建立了能快速、同时检测两种类病毒的RT-PCR方法。

Structural Engineering of Hierarchical Magnetic/Carbon Nanocomposites via In Situ Growth for High-Efficient Electromagnetic Wave Absorption

Materials exhibiting high-performance electromagnetic wave absorption have garnered considerable scientific and technological attention,yet encounter significant challenges.Developing new materials and innovative structural design concepts is crucial for expanding the application field of electromagnetic wave absorption.Particularly,hierarchical structure engineering has emerged as a promising approach to enhance the physical and chemical properties of materials,providing immense potential for creating versatile electromagnetic wave absorption materials.Herein,an exceptional multi-dimensional hierarchical structure was meticulously devised,unleashing the full microwave attenuation capabilities through in situ growth,selfreduction,and multi-heterogeneous interface integration.The hierarchical structure features a three-dimensional carbon framework,where magnetic nanoparticles grow in situ on the carbon skeleton,creating a necklace-like structure.Furthermore,magnetic nanosheets assemble within this framework.Enhanced impedance matching was achieved by precisely adjusting component proportions,and intelligent integration of diverse interfaces bolstered dielectric polarization.The obtain Fe_(3)O_(4)-Fe nanoparticles/carbon nanofibers/Al-Fe_(3)O_(4)-Fe nanosheets composites demonstrated outstanding performance with a minimum reflection loss(RLmin)value of−59.3 dB and an effective absorption bandwidth(RL≤−10 dB)extending up to 5.6 GHz at 2.2 mm.These notable accomplishments offer fresh insights into the precision design of high-efficient electromagnetic wave absorption materials.

Exploring the Cation Regulation Mechanism for Interfacial Water Involved in the Hydrogen Evolution Reaction by In Situ Raman Spectroscopy

Interfacial water molecules are the most important participants in the hydrogen evolution reaction(HER).Hence,understanding the behavior and role that interfacial water plays will ultimately reveal the HER mechanism.Unfortunately,investigating interfacial water is extremely challenging owing to the interference caused by bulk water molecules and complexity of the interfacial environment.Here,the behaviors of interfacial water in different cationic electrolytes on Pd surfaces were investigated by the electrochemistry,in situ core-shell nanostructure enhanced Raman spectroscopy and theoretical simulation techniques.Direct spectral evidence reveals a red shift in the frequency and a decrease in the intensity of interfacial water as the potential is shifted in the positively direction.When comparing the different cation electrolyte systems at a given potential,the frequency of the interfacial water peak increases in the specified order:Li+<Na^(+)<K^(+)<Ca^(2+)<Sr^(2+).The structure of interfacial water was optimized by adjusting the radius,valence,and concentration of cation to form the two-H down structure.This unique interfacial water structure will improve the charge transfer efficiency between the water and electrode further enhancing the HER performance.Therefore,local cation tuning strategies can be used to improve the HER performance by optimizing the interfacial water structure.

Optimizing 3d spin polarization of CoOOH by in situ Mo doping for efficient oxygen evolution reaction

Transition-metal oxyhydroxides are attractive catalysts for oxygen evolution reactions(OERs).Further studies for developing transition-metal oxyhydroxide catalysts and understanding their catalytic mechanisms will benefit their quick transition to the next catalysts.Herein,Mo-doped CoOOH was designed as a high-performance model electrocatalyst with durability for 20 h at 10 mAcm−2.Additionally,it had an overpotential of 260 mV(glassy carbon)or 215 mV(nickel foam),which was 78 mV lower than that of IrO_(2)(338 mV).In situ,Raman spectroscopy revealed the transformation process of CoOOH.Calculations using the density functional theory showed that during OER,doped Mo increased the spin-up density of states and shrank the spin-down bandgap of the 3d orbits in the reconstructed CoOOH under the electrochemical activation process,which simultaneously optimized the adsorption and electron conduction of oxygen-related intermediates on Co sites and lowered the OER overpotentials.Our research provides new insights into the methodical planning of the creation of transition-metal oxyhydroxide OER catalysts.

Investigation of an electrode-driven hydrogen plasma method for in situ cleaning of tin-based contamination

To prolong the service life of optics,the feasibility of in situ cleaning of the multilayer mirror(MLM)of tin and its oxidized contamination was investigated using hydrogen plasma at different power levels.Granular tin-based contamination consisting of micro-and macroparticles was deposited on silicon via physical vapor deposition(PVD).The electrodedriven hydrogen plasma at different power levels was systematically diagnosed using a Langmuir probe and a retarding field ion energy analyzer(RFEA).Moreover,the magnitude of the self-biasing voltage was measured at different power levels,and the peak ion energy was corrected for the difference between the RFEA measurements and the self-biasing voltage(E_(RFEA)-eV_(self)).XPS analysis of O 1s and Sn 3d peaks demonstrated the chemical reduction process after 1 W cleaning.Analysis of surface and cross-section morphology revealed that holes emerged on the upper part of the macroparticles while its bottom remained smooth.Hills and folds appeared on the upper part of the microparticles,confirming the top-down cleaning mode with hydrogen plasma.This study provides an in situ electrode-driven hydrogen plasma etching process for tin-based contamination and will provide meaningful guidance for understanding the chemical mechanism of reduction and etching.

In situ forming injectable MSC-loaded GelMA hydrogels combined with PD for vascularized sweat gland regeneration

Dear Editor,Three dimensional(3D)bioprinted extracellular matrix(ECM)can be used to provide both biochemical and biophysical cues to direct mesenchymal stem cells(MSCs)differentiation,and then differentiated cells were isolated for implantation in vivo using surgical procedures.However,the reduced cell activity after cell isolation from 3D constructs and low cell retention in injured sites limit its application[1].Methacrylated gelatin(GelMA)hydrogel has the advantage of fast crosslinking,which could resemble complex architectures of tissue construct in vivo[2].Here,we adopted a noninvasive bioprinting procedure to imitate the regenerative microenvironment that could simultaneously direct the sweat gland(SG)and vascular differentiation from MSCs and ultimately promote the replacement of glandular tissue in situ(Fig.1a).

Application of multidisciplinary in situ simulation training in the treatment of acute ischemic stroke: a quality improvement project

BACKGROUND:Ischemic stroke refers to a disorder in the blood supply to a local area of brain tissue for various reasons and is characterized by high morbidity,mortality,and disability.Early reperfusion of brain tissue at risk of injury is crucial for the treatment of acute ischemic stroke.The purpose of this study was to evaluate comfort levels in managing acute stroke patients with hypoxemia who required endotracheal intubation after multidisciplinary in situ simulation training and to shorten the door-to-image time.METHODS:This quality improvement project utilized a comprehensive multidisciplinary in situ simulation exercise.A total of 53 participants completed the two-day in situ simulation training.The main outcome was the self-reported comfort levels of participants in managing acute stroke patients with hypoxemia requiring endotracheal intubation before and after simulation training.A 5-point Likert scale was used to measure participant comfort.A paired-sample t-test was used to compare the mean self-reported comfort scores of participants,as well as the endotracheal intubation time and door-to-image time on the fi rst and second days of in situ simulation training.The door-to-image time before and after the training was also recorded.RESULTS:The findings indicated that in situ simulation training could enhance participant comfort when managing acute stroke patients with hypoxemia who required endotracheal intubation and shorten door-to-image time.For the emergency management of hypoxemia or tracheal intubation,the mean post-training self-reported comfort score was signifi cantly higher than the mean pre-training comfort score(hypoxemia:4.53±0.64 vs.3.62±0.69,t=-11.046,P<0.001;tracheal intubation:3.98±0.72 vs.3.43±0.72,t=-6.940,P<0.001).We also observed a decrease in the tracheal intubation and door-to-image time and a decreasing trend in the door-to-image time,which continued after the training.CONCLUSION:Our study demonstrates that the implementation of in situ simulation training in a clinical environment with a multidisciplinary approach may improve the ability and confi dence of stroke team members,optimize the fi rst-aid process,and eff ectively shorten the door-to-image time of stroke patients with emergency complications.