利用流式细胞仪检测人外周血NK细胞毒性方法的建立

目的利用流式细胞仪(FACS)技术检测人外周血NK细胞的毒性。方法首先将pEGFP-N1质粒转染人NK细胞的天然靶细胞K562,经过G418筛选后进一步单克隆化,得到稳定、均一表达绿色荧光蛋白的K562细胞。取对数期的K562-EGFP细胞与人外周血单个核细胞分别按5∶1、10∶1、20∶1、40∶1的比例混合,分别孵育0.5、1、2、4 h后用碘化丙啶(PI)标记,用流式细胞仪分析呈红、绿色荧光的细胞数,最后计算NK细胞的杀伤效率。结果0.5、1、2、4 h均能得到明显的杀伤效果,而且以2h的杀伤率最高。此时杀伤率与传统乳酸脱氢酶法(LDH)具有显著相关性(γ=0.997,P=0.003),而且显示出更强的敏感性。结论利用流式细胞仪检测NK细胞毒活性的方法可作为传统51Cr释放法、LDH法的一个补充,而且具有经济、快速和敏感的特点。     

流式细胞术检测NK细胞活性在获得性噬血细胞性淋巴组织细胞增多症诊断中的应用

本研究目的旨在建立一种准确、稳定的NK细胞杀伤活性检测方法,应用于获得性噬血细胞性淋巴组织细胞增多症诊断。21例疑似患者及20例健康对照者纳入本研究。根据HLH-2004标准将疑似患者分为确诊组和排除组,将pEGFP—N1质粒转染人NK细胞的天然靶细胞K562,经过G418筛选后进一步单克隆,得到稳定、均一表达绿色荧光蛋白的EGFP—K562细胞,将EGFP—K562细胞与人外周血单个核细胞按10：1的比例混合孵育2小时后,用碘化丙啶（PI）标记,用流式细胞仪分析呈红、绿色荧光的细胞数,最后计算NK细胞的杀伤效率;同时采用LDH释放法测定外周血NK细胞对单纯K562细胞的杀伤活性,并进行比较。结果表明：获得了稳定表达绿色荧光蛋白的EGFP—K562细胞;流式细胞仪检测表明健康人NK细胞杀伤率与病人组之间存在明显差异,流式细胞仪技术检测NK细胞杀伤活性与传统LDH释放法检测NK细胞杀伤活性具有显著相关性。结论：EGFP—K562细胞株作为靶细胞用于流式细胞术检测NK细胞杀伤活性,无需预先染色和标记靶细胞,操作简便、省时、稳定、重复性高,可应广泛应用于获得性噬血细胞性淋巴组织细胞增多症诊断。     

表达增强型绿色荧光蛋白K562细胞株的建立及其检测自然杀伤细胞活性的效果评价

目的建立一种准确、稳定的自然杀伤细胞(NK)活性检测方法。方法通过电穿孔法将含增强型绿色荧光蛋白(EGFP)的质粒转入K562细胞,经G418筛选后进行克隆,获得EGFP阳性表达细胞株;采用流式细胞仪和乳酸脱氢酶法检测10名健康人外周血NK细胞活性。结果通过基因转染的方法得到1株稳定表达EGFP的K562细胞,转染EGFP对细胞生长无影响,10名健康人外周血NK细胞对转染EGFP的细胞及未转染细胞的杀伤活性无差异(P>0.05);表达EGFP的K562细胞在效靶比为40∶1时,用流式细胞仪检测培养2和4h时的NK细胞杀伤率分别为(21.5±1.3)%和(23.7±1.5)%,二者无差异(P>0.05);而用乳酸脱氢酶法测定培养4h的NK细胞对K562和EGFP-K562的杀伤率[分别为(23.6±2.3)%和(23.7±2.3)%]均高于2h的杀伤率[(18.0±1.1)%和(18.2±1.3)%,P<0.05]。结论成功建立了表达EGFP的K562细胞株,将该细胞株作为靶细胞用于流式细胞术检测NK细胞活性时,无需预先染色和标记靶细胞,且操作简便、省时、稳定、可靠。     

姜黄素提高白血病KG1a细胞对同种异体NK细胞杀伤的敏感性及其分子机制探讨

目的：研究姜黄素作用前后白血病KG1a细胞对同种异体自然杀伤（NK）细胞杀伤的敏感性变化,并初步探讨其分子机制。方法：以对同种异体NK细胞杀伤高度敏感的白血病K562细胞为对照,乳酸脱氢酶释放法检测KG1a细胞对同种异体NK细胞杀伤的敏感性,流式细胞仪检测细胞表面NKG2D配体（MICA/B、ULBP1、ULBP2、ULBP3）和HLA-Ⅰ类分子蛋白表达,逆转录PCR法检测细胞表面NKG2D配体和HLA-Ⅰ基因mRNA的表达水平。CCK-8法检测姜黄素对KG1a细胞半数抑制量（IC50）,以此含量作用KG1a细胞,再次乳酸脱氢酶释放法检测姜黄素作用后的KG1a对同种异体NK细胞杀伤的敏感性,流式细胞仪检测姜黄素作用后的KG1a细胞表面NKG2D配体和HLA-Ⅰ类分子蛋白表达水平。结果：与K562细胞比较,KG1a细胞对同种异体NK细胞杀伤呈低度敏感,KG1a细胞表面NKG2D各配体蛋白表达阳性率均明显低于K562细胞（P〈0.01）,HLA-Ⅰ蛋白表达阳性率则明显高于K562细胞（P〈0.01）;在mRNA水平两种白血病细胞表面均有NKG2D配体和HLA-Ⅰ基因表达。当效靶比为5∶1、10∶1、20∶1时,姜黄素作用后NK细胞对KG1a细胞杀伤率明显高于作用前（P〈0.01）;作用后KG1a细胞各NKG2D配体表达率均明显升高（P〈0.05或0.01）,HLA-Ⅰ类分子无明显变化（P〉0.05）。结论：姜黄素能提高KG1a细胞对同种异体NK细胞杀伤的敏感性,其机制可能与姜黄素提高KG1a细胞表面NKG2D配体的表达有关。     

基于CD107a标记流式细胞仪检测NK细胞毒性方法的建立

目的建立基于CD107a标记、利用流式细胞仪检测NK细胞毒性的方法。方法首先将外周血单个核细胞（PBMCs）与K562细胞以3：1比例混合,2h后加入莫能菌素,1．5h后加入CD107a和CD56标记抗体,流式细胞仪分析CD107a阳性细胞的频率。其次观察CD107a抗体孵育时间、不同效靶比例对CD107a阳性细胞检测的影响。最后观察该方法与传统LDH释放法检测NK细胞毒活性的一致性。结果NK细胞活化后可在其表面检测到高水平表达的CD107a分子,效靶细胞孵育结束后加入CD107a抗体可降低检测背景,低效靶比例同样具有检测敏感性。该方法与LDH释放法的检测结果一致。结论利用CD107a抗体标记检测NK细胞毒活性的方法具有快速、敏感、所需效应细胞少的优点。     

树突状细胞对自体自然杀伤细胞体外扩增及功能的影响

本研究探讨树突状细胞（DC）对自体自然杀伤（NK）细胞体外扩增和功能的影响及其机制。用干细胞培养液（SCGM）在37℃、5％CO2、饱和湿度培养条件下以rhIL-2体外扩增NK细胞10天后，将自体DC与NK细胞以5：1（5：1组）和1：1（1：1组）的比例混合继续培养。14、21天时计算5：1组、1：1组和对照组的NK细胞扩增倍数，流式细胞术测定细胞表面CD3、CD56／16的表达，MTT法检测NK细胞的功能，ELISA法检测各培养组上清液中TNF-α和IL-12p70的含量。结果表明：14天时5：1、1：1组和对照组的扩增倍数分别为29．25±4．01、21．23±2．91和16．26±1．58，3组间比较差别有统计学意义（P〈0．05）。同组不同时间比较，14天时扩增倍数最高（P〈0．05）。14天时3组CD3^-、CD56／16^＋表达率分别为（64.6±7.8）％、（50．6±8.7）％和（34．8±5．1）％，5：1组表达率最高（P〈0．05）。14天时3组对K562细胞的杀伤率分别为（87.4±6．8）％、（75.4±6．3）％和（63．7±3.8）％，5：1组杀伤活性明显高于其它2组（P〈0．05）。5：1组上清液中TNF—α和IL-12p70的含量均分别高于其它各组（P均〈0．05）。结论：DC与NK细胞混合培养，能以比例依赖的方式增加NK细胞的扩增倍数，增强NK细胞的功能。DC增加NK细胞的扩增倍数与DC分泌的IL-12有关，NK细胞功能增强与NK细胞分泌的TNF—α增高有关。     

同种异体NK细胞对AML细胞体外杀伤活性及初步机制

目的探讨同种异体自然杀伤细胞(NK细胞)对急性髓系白血病(AML)细胞株KG1a的杀伤活性及其分子机制。方法以NK敏感的K562细胞株为对照,免疫磁珠法分离5例健康志愿者NK细胞,乳酸脱氢酶(LDH)释放法测定在不同效靶比时NK细胞对KG1a细胞的杀伤活性,并用流式细胞仪检测KG1a细胞表面人类白细胞抗原I(HLA-I)类分子和NKG2D的配体主要组织相容性复合体(MIC)A/B、ULBP1-3的表达情况。结果效靶比1∶1、5∶1、10∶1、20∶1时NK细胞杀伤K562和KG1a细胞的活性不同,NK细胞对KG1a细胞的杀伤活性较K562细胞明显减低(P<0.01);K562细胞低表达HLA-I类分子,高表达NKG2D配体MIC可溶性Ⅰ类分子相关蛋白A(MICA)、MICA有亲源性的蛋白(MICB)、ULBP1、ULBP2、ULBP3,而KG1a细胞高表达HLA-I类分子,低表达NKG2D配体MICA、MICB、ULBP1、ULBP2、ULBP3。结论同种异体NK细胞对AML细胞株KG1a的杀伤率低于K562细胞;其杀伤率不同的分子基础与其分子表面配体表达差异和HLA-I分子表达率不同有关。     

LDH释放法检测肿瘤患者PBMC和CIK细胞的细胞毒活性

目的建立检测细胞毒活性的乳酸脱氢酶（LDH）测定法，了解肿瘤患者外周血单个核细胞（PBMC）和细胞因子诱导的杀伤细胞（CIK）的抗肿瘤活性，为肿瘤的免疫细胞治疗提供参考。方法采集肿瘤患者和健康成人外周血，常规分离PBMC，体外进行CIK的培养，用LDH释放法分别检测PBMC和CIK对肿瘤细胞K562的细胞毒活性。结果肿瘤患者PBMC对K562的杀伤活力比正常人显著降低（P〈0．05）。经多种细胞因子诱导培养7～10d后，健康人和肿瘤患者的CIK对K562的杀伤活力都有显著提高（P〈0．01），肿瘤患者CIK杀伤活力接近于健康人。结论肿瘤患者PBMC细胞毒活性显著降低，但经诱导培养成CIK后则显著提高。用LDH释放法检测肿瘤病人CIK细胞毒活力，有助于CIK疗法适应病人的选择及个体疗效观察。     

K562细胞外泌体诱导特异性CTL生成的研究

为了研究人白血病细胞株K562细胞分泌的外泌体（exosomes）及K562细胞总RNA刺激人树突状细胞（DC）分泌的外泌体能否诱导出K562细胞特异性CTL，采用四步离心法提取K562细胞及K562细胞总RNA刺激人树突状细胞的培养上清液中的外泌体，并诱导CTL生成，以四甲基偶氮唑蓝（MTT）法检测CTL对K562细胞的杀伤作用。结果显示：K562细胞外泌体和K562细胞RNA刺激的DC和外泌体均能明显促进对K562细胞的杀伤活性，与未经外泌体作用的对照组相比有显著性差异（P〈0．05）。外泌体作用后对K562细胞的杀伤作用明显比对HL-60细胞的杀伤作用强，其差异有显著性（P〈0．05）。结论：K562细胞外泌体能够诱导出特异性抗白血病细胞的免疫反应。     

K562和K562/AO2细胞HLA I类分子与MICA/B的表达及其对NK细胞杀伤活性的影响

本研究探讨K562亲本细胞株及多药耐药细胞株K562/AO2细胞表面HLAI类分子和MHCI类链相关分子(MICA/B)的表达及其对NK细胞杀伤活性的影响。用流式细胞仪检测K562和K562/AO2细胞株HLAI类分子和MICA/B的表达情况;LDH释放法测定3例健康人NK细胞在不同效靶比时对K562和K562/AO2细胞的杀伤活性;效靶比10∶1时,用抗HLAI类分子单克隆抗体(W6/32)和抗MICA/B单克隆抗体(BAMO-1)分别封闭MHCⅠ类分子和MICA/B,观察NK细胞杀伤K562及K562/AO2细胞活性的变化。结果表明:K562和K562/AO2细胞均不表达HLAI类分子,K562细胞的MICA/B表达较K562/AO2明显增高(P<0.01)。效靶比5∶1、10∶1、20∶1、30∶1时NK细胞杀伤K562和K562/AO2细胞的活性分别为(29.32±0.12)%、(45.33±0.78)%、(58.37±0.87)%、(72.37±0.96)和(12.47±0.91)%、(24.36±1.11)%、(33.29±1.03)%、(53.87±1.27)%,各效靶比时NK细胞对K562细胞的杀伤活性较K562/AO2细胞明显增强(P=0.000);效靶比10∶1时W6/32不抑制NK细胞杀伤K562和K562/AO2细胞的活性,BAMO-1能明显抑制NK细胞杀伤K562和K562/AO2细胞的活性。结论:MICA/B在K562和K562/AO2细胞表达的差异与NK细胞的杀伤活性有关;NK细胞对K562/AO2细胞杀伤活性降低与HLAI类分子无关;MICA/B在多药耐药肿瘤细胞的低表达导致耐药肿瘤细胞对NK细胞杀伤敏感性下降。     

Determination of creatine kinase isoenzyme MB activity in serum using immunological inhibition of creatine kinase M subunit activity. Activity kinetics and diagnostic significance in myocardial infarction.

This is a new method for the determination of creatine kinase isoenzyme MB activity in serum. The method uses direct activity measurement of creatine kinase B subunit activity after blocking of CK-M subunit activity by inhibiting antibodies. The test takes no longer than 15 min. The method yields an intra-serial C.V. of 2.0-12.9%, and a C.V. from day to day of 5.5%. The detection limit is 3.4 U/l creatine kinase MB. In the 95 cases with proven myocardial infarction several types of creatine kinase MB activity kinetics could be determined. The percentage of creatine kinase MB of peak CK-total is 6-25%, with a mean of 11.1%. The amount of creatine kinase MB with respect to total CK activity after reinfarction is higher than the amount after initial infarction.     

俯卧位通气对高海拔地区肺复张治疗无效急性呼吸窘迫综合征患者氧合的影响

目的 探讨俯卧位通气对高海拔地区肺复张术(RM)治疗无效急性呼吸窘迫综合征(ARDS)患者的治疗作用.方法 从海拔2260m的地区医院筛选RM治疗无效的41例ARDS患者[平均氧合指数( PaO2/FiO2)较RM前升高＜20％视为RM无效],依不同病因分为肺内源性ARDS组(ARDSp组)和肺外源性ARDS组(ARDSexp组),每组再按信封法随机分为俯卧位组和仰卧位组,即ARDSp俯卧位组(11例)、ARDSp仰卧位组(9例)、ARDSexp俯卧位组(10例)、ARDSexp仰卧位组(11例).在通气前及通气1、2、3、4h监测动脉血氧分压( PaO2)、PaO2/FiO2、静态顺应性(Cst)、气道阻力(Raw)的变化.结果 通气lh时,ARDSexp俯卧位组PaO2/FiO2( mm Hg,l mm Hg=0.133 kPa)即较通气前显著升高(157.4±40.6比129.3±48.7,P＜0.05),并随通气时间延长呈持续增高趋势,4h达峰值(219.1 ±41.1);且ARDSexp俯卧位组通气3h内PaO2/FiO2较其他3组显著增高,另3组间则差异无统计学意义.ARDSp俯卧位组、ARDSexp俯卧位组通气4h时PaO2/FiO2均较相应仰卧位组显著增高(208.8±39.7比127.4±47.1,219.1±41.1比124.9±50.8,均P＜0.05).4组通气前后Cst无显著改变,各组间差异也无统计学意义.ARDSp俯卧位组通气4h时Raw(cmH2O·L-1·s-1)较通气前显著降低(6.8±1.7比10.7±1.8,P＜0.05),且明显低于其他3组;其他3组各时间点Raw组内及组间比较差异均无统计学意义.结论 俯卧位通气作为ARDS机械通气重要策略之一,可以改善RM无效高原ARDS患者的氧合,为抢救患者赢得宝贵的时间.     

Digital imaging among medical facilities

The Department of Veterans Affairs (VA) in the USA operates a network of 172 medical centres which all utilize a hospital information system (HIS) which has been developed and is currently maintained by the VA. During the past several years, an image management and communication module has been developed, installed and clinically utilized at the Washington DC and Maryland VA Medical Centres. This image management and communication system, referred to as the decentralized hospital computer program (DHCP) imaging system, is fully integrated with a commercial picture archiving and communication system (PACS). The system is utilized to capture, archive, and display all images generated within the hospital including radiology, nuclear medicine, pathology, endoscopy, bronchoscopy, and dermatology, intraoperative photographs, ECG data, and a limited number of paper documents. The ultimate goal of the project is to have all patient text and image data available at any clinical workstation to any authorized user anywhere within the network of medical centres. Clinical requirements for an imaging workstation include ease of use, rapid and reliable access to the complete set of patient information, and images which are of acceptable quality to meet the requirements of the user and the subspecialty. Patient confidentiality and data security must be safeguarded at all times. Integration of the images with the remainder of the patient's database was found to be critical to the success of the project. The experience at the Washington and Maryland facilities suggests that an imaging system that is successfully integrated with a hospital information system can provide substantial clinical and economic benefits both within and among medical centres. Clinical acceptance and utilization of the system has been excellent, particularly in diagnostic radiology where DHCP Imaging has been interfaced to a commercial PAC system. Based upon this initial experience, the VA has begun to deploy the system throughout its large network of medical centres.     

Myocardial elastography at both high temporal and spatial resolution for the detection of infarcts

Myocardial elastography is a novel method for noninvasively assessing regional myocardial function, with the advantages of high spatial and temporal resolution and high signal-to-noise ratio (SNR). In this paper, in-vivo experiments were performed in anesthetized normal and infarcted mice (one day after left anterior descending coronary artery [LAD] ligation) using a high-resolution (30 MHz) ultrasound system (Vevo 770, VisualSonics Inc., Toronto, ON, Canada). Radiofrequency (RF) signals of the left ventricle (LV) in longitudinal (long-axis) view and the associated electrocardiogram (ECG) were simultaneously acquired. Using a retrospective ECG gating technique, 2-D full field-of-view RF frames were acquired at an extremely high frame rate (8 kHz) that resulted in high-quality incremental displacement and strain estimation of the myocardium. The incremental results were further accumulated to obtain the cumulative displacements and strains. Two-dimensional and M-mode displacement images and strain images (elastograms), as well as displacement and strain profiles as a function of time, were compared between normal and infarcted mice. Incremental results clearly depicted cardiac events including LV contraction, LV relaxation and isovolumetric phases in both normal and infarcted mice, and also evidently indicated reduced motion and deformation in the infarcted myocardium. The elastograms indicated that the infarcted regions underwent thinning during systole rather than thickening, as in the normal case. The cumulative elastograms were found to have higher elastographic SNR (SNR(e)) than the incremental elastograms (e.g., 10.6 vs. 4.7 in a normal myocardium, and 6.0 vs. 2.4 in an infarcted myocardium). Finally, preliminary statistical results from nine normal (m = 9) and seven infarcted (n = 7) mice indicated the capability of the cumulative strain in differentiating infracted from normal myocardia. In conclusion, myocardial elastography could provide regional strain information at simultaneously high temporal (>/=0.125 ms) and spatial ( approximately 55 microm) resolution as well as high precision ( approximately 0.05 microm displacement). This technique was thus capable of accurately characterizing normal myocardial function throughout an entire cardiac cycle, at the same high resolution, and detecting and localizing myocardial infarction in vivo.     

手转胎头术失败的原因与分娩结局临床分析

目的 探讨手转胎头术失败的原因与分娩结局.方法 选择2008年1月至2010年12月于我院住院分娩的持续性枕横位、枕后位产妇198例,根据行手转胎头术后结果分为成功组126例、失败组72例.比较两组分娩结局,对比分析失败原因.结果 失败组胎儿体质量≥3500 g的发生率[76.4％(55/72)]明显高于成功组[31.7％(40/126)],差异有统计学意义(x2=30.177,P=0.001)、失败组宫缩乏力发生率[58.3％(42/72)]高于成功组[38.1％ (48/126)],差异有统计学意义(x2=7.569,P=0.006)、失败组骨盆临界或轻度狭窄发生率[38.9％ (28/72)]高于成功组[23.8％(30/126)],差异有统计学意义(x2 =5.030,P=0.002)、失败组手转胎头时机不当(宫口开大＜6 cm、胎头位于坐骨棘上及宫口开大8～10 cm、胎头位于坐骨棘下≥2 cm)发生率[61.1％(44/72)]高于成功组[38.9％(49/126)],差异有统计学意义(x2=9.084,P=0.003).失败组母儿并发症(产后出血、产褥病率、胎儿窘迫、新生儿窒息)发生率高于成功组(x2 =9.586,P=0.002、x2=9.334,P=0.002、x2=5.910,P=0.015、x2=5.240,P=0.022)、失败组剖宫产发生率[72.2％(52/72)]明显高于成功组[34.1 ％(43/126),x2=26.641,P=0.001)].结论 手转胎头术能使难产变顺产,降低剖宫产率,减少母儿并发症,但须积极预防、处理导致手转胎头术失败的原因,对矫正失败后继续矫正及试产应慎重.     

Clinical Pharmacokinetics of Morphine

Morphine, the most widely used mu-opioid analgesic for acute and chronic pain, is the standard against which new analgesics are measured. A thorough understanding of the pharmacokinetics of morphine is required in order to safely and effectively use this analgesic in a wide variety of patients with different levels of organ function. A MEDLINE search was conducted to identify literature published between 1966 and January 2002 relevant to the pharmacokinetics of morphine. These publications were reviewed and the literature summarized regarding unique and clinically important elements of morphine disposition relative to its parenteral administration (including intravenous, intramuscular, subcutaneous, epidural and intrathecal administration), absorption profile (immediate release, controlled release, and sublingual/buccal, and rectal administration), distribution, and its metabolism/ excretion. Special populations, including infants, elderly, and those with renal/liver failure, have a unique morphine pharmacokinetic profile that must be taken into account in order to maximize analgesic efficacy and reduce the risk of adverse events.     

Evidence-Based Pain Management and Palliative Care in Issue Four for 2009 of The Cochrane Library

ABSTRACT

The Cochrane Library of Systematic Reviews is published quarterly. Issue 4 for 2009 contains 4027 complete reviews, 1906 protocols for reviews in production, and 11447 one-page summaries of systematic reviews published in the general medical literature. In addition, there are citations of 600,000 randomized controlled trials, and 12,200 cited papers in the Cochrane methodology register. The health technology assessment database contains over 7500 citations. This edition of the Library contains 90 new reviews, of which 19 have potential relevance for practitioners in pain and palliative medicine.     

Wie zuverlässig sind Angaben zu nichtentzündlichen ausgedehnten Schmerzen?

ZusammenfassungFragestellung Es wurde geprüft, wie sich der Differenziertheitsgrad zweier Schmerzmessmethoden auf Angaben zur Ausgedehntheit klinischer Schmerzen auswirkt. Zugleich wurde der Referenzzeitraum variiert, über den die Patienten berichten sollten.Methode Erfasst wurde der Einfluss zu Lasten der Befragungsdifferenziertheit durch den Vergleich zweier Körperschema-Bildvorlagen. Drei Referenzzeiträume (Schmerz aktuell, letzte Woche, letztes halbes Jahr) wurden vorgegeben.Ergebnisse Patienten mit ausgedehnten Schmerzen gaben bei differenzierter Befragung um so mehr Schmerzen an, je weiter die Schmerzen zurück lagen und je größer der Berichtszeitraum war. Patienten mit gelenknahen Schmerzen gaben bei hoch differenzierter Befragung weniger ausgedehnte Schmerzen in der Vergangenheit an als bei globaler Einschätzung. Patienten mit Rückenschmerzen berichteten bei differenzierter Befragung zum aktuellen Schmerz über weniger ausgedehnte Schmerzen als bei globaler Befragung.Schlussfolgerung Die Angaben zur Schmerzausdehnung variieren vor allem bei Patienten mit ausgedehnten Schmerzen in Abhängigkeit von der Differenziertheit der Befragung. In diesen Fällen ist die Wahrscheinlichkeit erhöht, dass sich die Beschwerdesymptomatik zumindest teilweise erst in der Reaktion auf die situativen Befragungsbedingungen konstituiert und daher nicht auf andere Befragungsbedingungen generalisiert werden kann.