循环肿瘤细胞水平与卵巢癌患者临床病理特征及生存率的关系研究

目的:研究循环肿瘤细胞(CTC)水平与卵巢癌患者临床病理特征及生存率的关系。方法:选择从2012年6月到2014年12月在我院就诊的卵巢癌患者94例记为观察组。根据所有患者CTC水平将其分成高CTC组(CTC水平≥7个/5mL)57例及低CTC组(CTC水平7个/5 m L)37例,另选同期在我院体检的卵巢良性囊肿者90例记为对照组,分析CTC阳性与患者病理特征的关系、CTC水平与患者生存率和复发转移的关系以及CTC水平与患者病理特征、生存率、复发转移率的相关性。结果:对照组90例患者均未检测出CTC阳性。观察组94例患者中检测出68例CTC阳性,总阳性率为72.34%;观察组患者临床分期为II期和III期的CTC阳性率均明显高于I期,有淋巴结转移的CTC阳性率明显高于无淋巴结转移(P0.05)。高CTC组的1年、2年、3年生存率与低CTC组相比,差异均无统计学意义(P0.05)。高CTC组的1年、2年、3年复发转移率均高于低CTC组(P0.05)。Spearman相关性分析结果显示,CTC水平与患者的临床分期、淋巴结转移以及复发转移率均呈正相关(P0.05),而与年龄、病理分型、病理分级及生存率均无明显相关性(P0.05)。结论:卵巢癌患者临床分期为Ⅱ期和Ⅲ期和有淋巴结转移的CTC阳性率较高,且CTC水平与复发转移率关系密切,临床通过检测CTC水平,可能有助于评估患者的预后。     

多西他赛或紫杉醇联合奈达铂在宫颈癌辅助化疗中的疗效及安全性分析

目的:观察和比较多西他赛或紫杉醇联合奈达铂辅助化疗治疗宫颈癌患者的临床疗效及其安全性。方法:选择45例采用多西他赛联合奈达铂化疗的宫颈癌患者为观察组及45例同期采用紫杉醇联合奈达铂化疗的宫颈癌患者作为对照组,两组均行手术治疗,且术前接受辅助化疗。对比两组临床疗效、手术时间和术后病理状况及不良反应的发生情况。结果:化疗后,观察组临床有效率高于对照组(62.22%vs.55.56%),但组间比较差异无统计学意义(P0.05)。化疗期间,观察组恶心、呕吐、腹痛、腹泻、白细胞、中性粒细胞减少、血红蛋白和血小板减少的发生率均低于对照组,但组间比较差异均无统计学意义(P0.05);观察组神经毒性发生率明显低于对照组,组间比较差异有统计学意义(P0.05)。化疗后,行手术治疗,观察组手术时间低于对照组,但组间比较差异无统计学意义(P0.05);手术后,观察组盆腔淋巴结转移率和宫旁浸润率低于对照组,但组间比较差异无统计学意义(P0.05)。结论:多西他赛联合奈达铂辅助化疗宫颈癌的疗效与紫杉醇联合奈达铂相当,且神经毒性、骨髓抑制方面的发生率明显降低,是临床低毒性且有效的宫颈癌术前新辅助化疗方案。     

血清抑制素B评估成年隐睾患者术后睾丸功能恢复的临床价值

目的:探讨血清抑制素B(InhB)用于评价成年隐睾患者术后睾丸功能恢复的临床价值。方法:收集2011年10月~2015年9月于我科就诊的138例成年隐睾患者作为观察组,另收集100例健康志愿者作为正常对照组。用双抗体夹心酶联免疫吸附法检测观察组患者术前、术后30天和90天及正常对照组的血清InhB水平,并参照WHO人类精液检查与处理实验室手册(第五版)方法进行精液常规检查。结果:与正常对照组比较,观察组患者术前、术后30天和90天血清InhB水平、精子活率、精子密度均明显降低,差异均有统计学意义(P0.05)。观察组术后30天血清InhB水平、精子活率、精子密度与术前比较差异均无统计学意义(P0.05),观察组术后90天血清InhB水平、精子活率、精子密度明显高于术前,差异均有统计学意义(P0.05)。观察组组术前、术后30天和90天的精液量与正常对照组对比差异均无统计学意义(P0.05)。观察组术前、术后30天和90天及正常对照组血清InhB水平与精子浓度均呈正相关(观察组术前:r=0.81,P0.05,观察组术后30天:r=0.78,P0.05,观察组术后90天:r=0.84,P0.05,正常对照组:r=0.77,P0.05)。结论:血清InhB用于评估成年隐睾患者术后睾丸功能恢复具有一定的临床应用价值。     

手术前后注射帕瑞昔布钠对脑膜瘤切除术后的镇痛效果评价

目的:比较手术前后注射帕瑞昔布钠对脑膜瘤切除术后的镇痛效果。方法:收集我院收治的120例行脑膜瘤切除术患者,随机分为术前组、术后组以及对照组,每组40例。术前组在麻醉前15 min静注帕瑞昔布钠40 mg所有患者行麻醉诱导(咪达唑仑+异丙酚+罗库溴铵+舒芬太尼)静脉注射,维持麻醉采用异丙酚+瑞芬太尼静脉泵注,行脑膜瘤切除术术毕根据患者具体情况停用异丙酚、瑞芬太尼术后组患者在手术结束前15 min静注帕瑞昔布钠40 mg,对照组患者在手术结束前15 min静注生理盐水2mL。观察并比较三组患者各时间点的疼痛视觉模拟评分(VAS)、简易智能状态量表(MMSE)评分、血清皮质醇(cortisol)水平以及患者不良反应发生率。结果:与对照组相比术前组、术后组患者术后1 h的VAS评分水平较低差异具有统计学意义(P0.05);与术后组相比术前组患者的术后1 h的VAS评分水平较低差异具有统计学意义(P0.05);术后三组患者的MMSE评分与术前相比均下降(P0.05),与对照组相比术前组患者的术后1 h、术后24h MMSE评分较高差异具有统计学意义(P0.05);与对照组相比术前组、术后组患者术后1 h以及术后24 h的皮质醇水平较低差异具有统计学意义(P0.05);与术后组相比术前组患者的皮质醇水平较低差异具有统计学意义(P0.05);三组患者的副作用发生率相比无明显差异(P0.05)。结论:术前注射帕瑞昔布钠的镇痛效果较术后注射更好术后认知功能恢复较快。     

伊立替康联合卡铂辅助化疗对宫颈癌患者血清叶酸和CA125水平的影响

目的:探究伊立替康联合卡铂辅助化疗对宫颈癌患者血清叶酸、CA125及临床疗效的影响。方法:选择我院收治的宫颈癌患者41例,随机分成实验组和对照组。对照组19例予广泛全子宫切除及盆腔淋巴结清扫术;实验组22例在行全子宫切除及盆腔淋巴结清扫术前予卡铂和伊立替康治疗。对比两组的临床疗效以及治疗前后患者血清叶酸及CA125水平的变化。结果:实验组患者的有效率(81.8%)高于对照组(47.4%),其差异有统计学意义(P0.05);治疗后,两组患者的血清CA125水平均降低、叶酸水平均升高(P0.05),其差异有统计学意义;与对照组相比,实验组叶酸水平较高、血清CA125水平较低(P0.05),差异有统计学意义。结论:伊立替康联合卡铂辅助化疗对宫颈癌患者有较好的临床疗效,推测其机制可能与降低患者CA125水平及提高叶酸水平有关。     

PTVE 与TIPS治疗肝硬化门静脉高压合并食管胃底静脉曲张破裂出血 的疗效比较

目的:研究经皮胃底曲张静脉栓塞术(PTVE)和经颈静脉肝内门体分流术(TIPS)治疗肝硬化门静脉高压合并食管胃底静脉曲张破裂出血的临床疗效,为临床治疗提供依据。方法:选取2001年4月到2015年4月我院肝硬化门静脉高压合并食管胃底静脉曲张破裂患者169例,根据手术方式分为PTVE组(行PTVE治疗)141例和TIPS组(行TIPS治疗)28例,比较两组术前、术后门静脉压力,术前、术后3个月、6个月以及1年两组Child-Pugh评分、白蛋白以及直接胆红素,并比较两组再出血和肝性脑病发生率。结果:TIPS组术后门静脉压力较术前显著降低,比较差异具有统计学意义(P0.05),术后PTVE组及TIPS组组间比较差异具有统计学意义(P0.05);两组术前和术后各时间直接胆红素无统计学意义(P0.05);PTVE组术后1年白蛋白水平显著升高,与术前和TIPS组比较差异具有统计学意义(P0.05),TIPS组术后白蛋白有所升高,但各时间比较差异无统计学意义(P0.05),PTVE组术后各时间Child-Pugh评分较术前明显改善,比较差异具有统计学意义(P0.05),TIPS组术后3个月和术后6个月Child-Pugh评分较术前明显改善,比较差异具有统计学意义(P0.05);两组术后再出血发生率比较无统计学意义(P0.05),PTVE组肝性脑病发生率显著低于TIPS组,比较差异具有统计学意义(P0.05)。结论:PTVE和TIPS治疗肝硬化门静脉高压合并食管胃底静脉曲张破裂出血效果相当,TIPS能显著降低门静脉压,PTVE能降低肝性脑病的发生率,改善患者Child-Pugh评分。     

TP新辅助化疗方案治疗Ib-Ⅱa期局部晚期宫颈癌的临床效果观察

目的:观察并探讨宫颈癌术前行TP与PF两种新辅助化疗方案的临床效果,优选新辅助化疗方案。方法:将2008年1月-2010年12月间入选的91例Ib-Ⅱa宫颈癌患者随机分为A(n=46)、B(n=45)两组,A组患者子宫切除术前予TP新辅助化疗方案,B组予PF方案,各2个周期,对比化疗结束后两组临床疗效、化疗期间药物毒性反应及术后2年内宫颈癌复发率。结果:①A、B两组总体有效率分别为89.1%、82.2%,差异不具有统计学意义(P0.05),但前者化疗后瘤体直径明显小于B组(P0.05);②两组药物毒性反应构成不具有统计学差异(P0.05),但A组胃肠道反应与肝脏毒性发生率明显低于B组(P0.05);③A、B两组组术后2年内宫颈癌复发率分别为2.2%、11.9%,复发率无统计学差异(P0.05)。结论:宫颈癌术前TP新辅助化疗方案较之BIP方案具有比较优势,更具备临床推广价值。     

脑胶质瘤患者手术前后睾酮水平变化及临床意义

目的:研究脑胶质瘤患者手术前后睾酮水平变化,并分析其临床意义。方法:选取2010年2月到2015年6月我院脑胶质瘤患者80例为研究组,并选择同期颅脑良性肿瘤患者80例为对照组,两组均给予手术治疗,检测术前和术后3天、术后1周患者睾酮水平。结果:研究组术后各病理类型睾酮水平较治疗前显著降低(P0.05),但不同病理类型之间睾酮水平比较无统计学意义(P0.05);研究组术前睾酮水平显著高于对照组,比较差异具有统计学意义(P0.05);术后3天、术后一周研究组睾酮水平与术前比较显著降低,差异具有统计学意义(P0.05);术后1周研究组睾酮水平与术后3天比较无统计学意义(P0.05);对照组术前和术后各时间睾酮水平比较无统计学意义(P0.05)。结论:脑胶质瘤患者会出现睾酮水平紊乱,当睾酮水平明显增高时应警惕脑胶质瘤的可能性。     

腹腔镜与开腹宫颈癌根治术对患者血清IL-4, IL-10, TNF-α及IFN-γ水平的影响

目的:探讨腹腔镜与开腹宫颈癌根治术对患者血清白细胞介素-4(IL-4)、白细胞介素-10(IL-10)、肿瘤坏死因子-α(TNF-a)、γ-干扰素(IFN-γ)水平的影响。方法:选取2014年3月至2016年9月在我院接受治疗的64例宫颈癌患者作为研究对象,通过随机方式分为腹腔组和开腹组。比较两组患者的手术时间、术中出血量、术后通气时间、治疗前后血清IL-4、IL-10、TNF-a、IFN-r水平的变化。结果:腹腔组患者的术中出血量和术后通气时间明显低于或短于开腹组(P0.05),而两组患者的手术时间比较差异无统计学意义(P0.05)。术前,两组患者的血清IL-4、IL-10、TNF-α及IFN-γ水平比较差异均无统计学意义(P0.05);术中,腹腔组的血清IL-4水平明显低于开腹组(P0.05),其余三组指标水平与开腹组比较差异无统计学意义(P0.05);术后1天和7天,腹腔组患者的血清IL-4、IL-10、TNF-α及IFN-γ水平均明显低于开腹组(P0.05)。结论:腹腔镜宫颈癌根治术在术中失血量更少,安全性更高,且对于患者术后的IL-4、IL-10、TNF-α和IFN-γ水平抑制效果更加明显,利于患者的术后恢复。     

应用改良快速顺序诱导快速康复外科全麻效果观察

目的:探讨改良快速顺序诱导用于快速康复外科全麻患者的临床效果。方法:随机选择我院2015年1月1日至2015年12月31日胃肠外科行腹腔镜下胃癌根治手术患者共计100例,随机分为观察组和对照组各50例,分别行改良快速顺序诱导和经典快速顺序诱导。并比较术前基本情况、术中情况、术后情况及免疫相关指标。结果:两组患者手术时间、手术方式、肿瘤分期方面分析比较,差异无统计学意义(P0.05);观察组术中出血量及补液量均少于对照组,差异具有统计学意义(P0.05)。观察组术后排气时间、排便时间、住院时间、进食时间均明显早于对照组,差异具有统计学意义(P0.05)。术前、术后1日、术后3日、出院前,Ig G差异均具有统计学意义(P0.05)。结论:改良快速顺序诱导用于快速康复外科全麻患者可加速患者术后的恢复,提升术后免疫力。     

用CTC荧光染色法检测添加肝素对牛精子体外获能的影响

本试验在TALP液中添加不同浓度(0、20、40、80μg.ml-1)的肝素对牛精子进行获能,用CTC染色检测精子获能状况,探讨肝素浓度及诱导时间(1、3、6、9h)对牛精子体外获能的影响。结果显示:随着肝素浓度的提高,精子获能效果增强,3 h达到最高,但添加80μg.ml-1的肝素精子的顶体反应率低于其他组。结果表明:在TALP液中增加高浓度的肝素对精子体外获能有促进作用,用高浓度的肝素获能处理3h后的精子进行体外受精可能会得到较好的效果。     

Comparison of the relative efficacy of an insect baiting method and selective media for diversity studies of Metarhizium species in the soil

Metarhizium are a commonly occurring group of entomopathogenic fungi normally found in soil. The most common methods to assess the diversity of Metarhizium species in soil are (i) the use of selective media and (ii) insect baiting using Galleria mellonella larvae. We compared the recovery efficiency from soil of four common species of Metarhizium (Metarhizium anisopliae, Metarhizium pingshaense, Metarhizium brunneum and Metarhizium robertsii) using these two methods. Firstly, we compared the number of colony forming units (CFU) produced in vitro when grown on two selective media, one containing chloramphenicol, thiabendazole and cycloheximidethe (CTC) and one based on the fungicide dodine (n-dodecylguanidine acetate) (DOD). Secondly, we artificially inoculated natural/non-sterile soil with the four fungal species at a rate of 2×10² and 2×10³ conidia g^?1of soil, baited with G. mellonella, and processed for evaluation using the selective media. The in vitro results showed that the greatest number of CFUs were recorded for M. brunneum. In contrast, when inoculated into soil, more G. mellonella larvae became infected by M. anisopliae. Finally, when using selective media, most CFUs recovered were for M. robertsii. The importance of our results in selecting a method to study the natural occurrence of Metarhizium in soil are discussed.     

Changes in calmodulin immunocytochemical localization associated with capacitation and acrosomal exocytosis of ram spermatozoa

The aim of this study was to determine the localization of calmodulin (CaM) in ram sperm and the possible changes during in vitro capacitation (CA) and the ionophore-induced acrosome reaction (AR). Likewise, changes in intracellular calcium levels ([Ca²⁺]_i) were also analysed by using flow cytometry. CA was induced in vitro in a medium containing BSA, CaCl₂, NaHCO₃, and AR by the addition of the calcium ionophore A23187. The acrosomal status was assessed by the chlortetracycline-fluorescence (CTC) assay. Flow cytometry (FC) analyses were performed by loading samples with Fluo-3 AM, that emits fluorescence at a high [Ca²⁺]_i, combined with propidium iodide (PI) that allowed us to discriminate sperm with/without an integral plasma membrane both with high/low [Ca²⁺]_i. Immunocytochemistry localized CaM to the flagellum, and some sperm also contained CaM in the head (equatorial and post-acrosomal regions). CA and AR resulted in a slight increase in the post-acrosomal labelling. The treatment of sperm with increasing concentrations of two CaM antagonists, W7 and calmidazolium (CZ), accounted for an increase in capacitated and acrosome-reacted CTC-sperm patterns. CZ induced a significant reduction in the content of three protein tyrosine-phosphorylated bands of approximately of 30, 40 and 45 kDa. However, W7 showed no significant effect at any of the studied concentrations. Neither of them significantly influenced protein serine and threonine phosphorylation. FC analysis revealed that the main subpopulation in the control samples contained 70% of the total sperm with integral plasma membrane and a medium [Ca²⁺]_i. After CA, 67.1% of the sperm preserved an integral membrane with a higher [Ca²⁺]_i. After AR, only 7.2% of the total sperm preserved intact membranes with a very high [Ca²⁺]_i. These results imply that CaM appears to be involved in ram sperm capacitation, and both treatments increased its localization in the post-acrosomal region.     

ViBiBa: Virtual BioBanking for the DETECT multicenter trial program - decentralized storage and processing

BackgroundLiquid Biopsy (LB) in the form of e.g., circulating tumor cells (CTCs) is a promising non-invasive approach to support current therapeutic cancer management. However, the proof of clinical utility of CTCs in informing therapeutic decision-making for e.g., breast cancer in clinical trials and associated translational research projects is facing the issues of low CTC positivity rates and low CTC numbers – even in the metastasized situation. To compensate for this dilemma, clinical CTC trials are designed as large multicenter endeavors with decentralized sample collection, processing and storage of products, making data management highly important to enable high-quality translational CTC research.AimIn the DETECT clinical CTC trials we aimed at developing a custom-made, browser-based virtual database to harmonize and organize both decentralized processing and storage of LB specimens and to enable the collection of clinically meaningful LB sample.MethodsViBiBa processes data from various sources, harmonizes the data and creates an easily searchable multilayered database.ResultsAn open-source virtual bio-banking web-application termed ViBiBa was created, which automatically processes data from multiple non-standardized sources. These data are automatically checked and merged into one centralized databank and are providing the opportunity to extract clinically relevant patient cohorts and CTC sample collections.SummaryViBiBa, which is a highly flexible tool that allows for decentralized sample storage of liquid biopsy specimens, facilitates a solution which promotes collaboration in a user-friendly, federalist and highly structured way. The source code is available under the MIT license from https://vibiba.com or https://github.com/asperciesl/ViBiBa     

UHRF1基因在乳腺癌循环肿瘤细胞中的表达研究

外周血液中乳腺循环癌肿瘤细胞的生物表征与转移性乳腺癌的严重程度密切相关,本研究的目的在于结合体外细胞实验探讨UHRF1基因对乳腺癌进展的意义。多重RNA原位分析乳腺癌循环肿瘤细胞(circulating tumor cells,CTCs)中UHRF1的表达;MTT法检测UHRF1基因转染对正常乳腺细胞增殖的影响;蛋白免疫印迹检测UHRF1基因转染对正常乳腺细胞中Bax蛋白和Bcl-2蛋白的影响;Caspase-3检测试剂盒检测正常乳腺细胞中Caspase-3活性;Transwell侵袭实验和划痕愈合实验检测UHRF1基因转染对正常乳腺细胞侵袭和迁移能力的影响。研究发现,UHRF1 RNA水平在乳腺癌循环肿瘤细胞中高表达;UHRF1基因增加正常乳腺细胞增殖率;UHRF1基因降低正常乳腺细胞中Caspase-3活性;UHRF1基因降低正常乳腺细胞中Bax蛋白的表达,增加Bcl-2蛋白的表达;UHRF1基因增强正常乳腺细胞侵袭和迁移能力。本研究初步说明,UHRF1可促进正常乳腺细胞增殖,抑制正常乳腺细胞凋亡,增强正常乳腺细胞侵袭和迁移能力。     

Detection and isolation of circulating tumor cells: Principles and methods

Efforts to improve the clinical management of several cancers include finding better methods for the quantitative and qualitative analysis of circulating tumor cells (CTCs). However, detection and isolation of CTCs from the blood circulation is not a trivial task given their scarcity and the lack of reliable markers to identify these cells. With a variety of emerging technologies, a thorough review of the exploited principles and techniques as well as the trends observed in the development of these technologies can assist researchers to recognize the potential improvements and alternative approaches. To help better understand the related biological concepts, a simplified framework explaining cancer formation and its spread to other organs as well as how CTCs contribute to this process has been presented first. Then, based on their basic working-principles, the existing methods for detection and isolation of CTCs have been classified and reviewed as nucleic acid-based, physical properties-based and antibody-based methods. The review of literature suggests that antibody-based methods, particularly in conjunction with a microfluidic lab-on-a-chip setting, offer the highest overall performance for detection and isolation of CTCs. Further biological and engineering-related research is required to improve the existing methods. These include finding more specific markers for CTCs as well as enhancing the throughput, sensitivity, and analytic functionality of current devices.     

Inhibition of autophagy of Cajal mesenchymal cells by gavage of tong bian decoction based on the rat model of chronic transit constipation

The objective of this research was to study the effect of tong bian decoction on colon transport function of interstitial cells of Cajal (ICC) in chronic transit constipation (CTC) and the inhibition of autophagy of ICC, so as to achieve the free movement of the bowels. In this research, the experimental rats were divided into normal group (NG) and model group (MG) by random method, and the rat model of CTC was constructed by subdivision circulatory increasing operation gavage method of rhubarb. After the successful establishment of the model, the rats were divided into normal group, MG, tong bian decoction gavage group, mosapride group and normal recovery group. Then, rats in the NG and the MG were killed at the same time, and rats in the tong bian decoction gavage group, mosapride group and normal recovery group were killed at the same time. In this study, the transport function of colon of rats in each group was detected by activated carbon method, and the number of fecal residues in the colon was observed. The mRNA expression of c-kit gene in intestinal tissue of rat was detected by real-time quantitative polymerase chain reaction (RT-qPCR). In addition, the changes of ICC in rats treated with different drugs were detected by immunohistochemical method. The results revealed that in the tong bian decoction gavage group, the water content in the feces of rats was remarkably increased (P < 0.05), the amount of residual feces in the colon was remarkably reduced (P < 0.01), the percentage of carbon powder propulsion in small intestine was remarkably increased (P < 0.01), the staining area of ICC positive cells in colon tissue was remarkably increased (P < 0.05), and the expression of c-kit mRNA was remarkably increased (P < 0.01). It can be concluded that the tong bian decoction could effectively enhance the colon transport function in the rat model of CTC. This laxative mechanism promotes the regeneration and repair ability of ICC by inhibiting the autophagy of ICC, and provides power for the large intestine, so as to achieve the free movement of the bowels. Therefore, the results of this study have certain guiding meaning for the treatment of CTC with traditional Chinese medicine.     

5S rRNA-recognition module of CTC family proteins and its evolution

The effects of amino acid replacements in the RNA-binding sites of homologous ribosomal proteins TL5 and L25 (members of the CTC family) on ability of these proteins to form stable complexes with ribosomal 5S RNA were studied. It was shown that even three simultaneous replacements of non-conserved amino acid residues by alanine in the RNA-binding site of TL5 did not result in noticeable decrease in stability of the TL5-5S rRNA complex. However, any replacement among five conserved residues in the RNA-binding site of TL5, as well as of L25 resulted in serious destabilization or complete impossibility of complex formation. These five residues form an RNA-recognition module in TL5 and L25. These residues are strictly conserved in proteins of the CTC family. However, there are several cases of natural replacements of these residues in TL5 and L25 homologs in Bacilli and Cyanobacteria, which are accompanied by certain changes in the CTC-binding site of 5S rRNAs of the corresponding organisms. CTC proteins and specific fragments of 5S rRNA of Enterococcus faecalis and Nostoc sp. were isolated, and their ability to form specific complexes was tested. It was found that these proteins formed specific complexes only with 5S rRNA of the same organism. This is an example of coevolution of the structures of two interacting macromolecules.     

Implications of cancer-associated systemic inflammation for biomarker studies

Highly sensitive molecular technologies provide new capacities for cancer biomarker research, but with sensitivity improvements marker specificity is significantly decreased, and too many false-positive results should disqualify the measurement from clinical use. Hence, of the thousands of potential cancer biomarkers only a few have found their way to clinical application. Differentiating false-positive results from true-positive (cancer-specific) results can indeed be difficult, if validation of a marker is performed against inadequate controls.     

Assessment of metabolic activity of single bacterial cells — new developments in microcolony and dehydrogenase assays

Abstract: Some bacteria lose culturability in natural environments but retain measurable metabolic activity and are thus considered viable. Several techniques have been proposed to determine the activity of nonculturable cells. Due to the considerable physiological heterogeneity of bacterial populations in the environment, it is imperative to apply methods which measure cellular activity at the single cell level. This review focuses on two promising methods: the microcolony assay and the respiration assay based on reduction of 5-cyano-2,3-ditolyl tetrazolium chloride (CTC). In the microcolony assay, viable cells are identified by their ability to perform a limited number of cell divisions and this approach is thus related to conventional culture techniques. Some recent methodological developments of the technique aiming at improving the incubation conditions and the detection of microcolonies are presented. Results obtained by the microcolony technique are used to introduce its advantages and limitations. The CTC-reduction assay determines a central cellular metabolic activity, but does not measure cell growth. Results of studies using this assay are presented, and it is emphasized that great care should be taken to optimize assay conditions for the studied organisms. Finally, the results obtained by different viability assays are compared. For a specific bacterium, several assays, addressing different aspects of cell metabolism, can provide comparable results suggesting that they provide meaningful viability estimates. On the other hand, the use of viability assays on complex indigenous populations may be ambiguous.