目的 探讨结直肠癌原发灶及转移淋巴结中胸腺嘧啶核苷酸合成酶(thymidylate synthase,TS)、Ki-67和p53表达差异性,并探究其对预测5-氟尿嘧啶(5-fluorouracil,5-FU)敏感性的价值.方法 选取有淋巴结转移的结直肠腺癌患者33例,采用免疫组织化学方法检测结直肠癌淋巴结转移灶及原发灶中TS、Ki-67和P53的表达情况.结果 TS高表达、p53和Ki-67在转移淋巴结的表达率显著高于原发灶;较好预后的即对5-FU化疗敏感的病例共7例,其中在转移淋巴结及原发癌中TS均为低表达率,p53、Ki-67为阴性表达.结论 转移淋巴结中的p53、TS、Ki-67的表达情况同原发灶相比有异质性变化,转移淋巴结TS表达情况可以预测结直肠癌患者对5-FU化疗敏感性,p53的表达情况或许可预测结直肠癌患者对5-FU的化疗敏感性,Ki-67表达与5-FU化疗敏感性的关系仍需要进一步研究.

目的 研究大肠癌原发灶和相应淋巴结转移灶中胸苷磷酸化酶(thymidine phosphorylase,TP)及血管内皮细胞生长因子(vascular endothelial growth factor,VEGF)的表达及与5-氟尿嘧啶(5-fluorouracil,5-FU)化疗敏感性的关系.方法 收集2010年1月至2012年3月间上海交通大学医学院附属新华医院崇明分院诊治的有淋巴结转移的大肠腺癌患者;通过免疫组织化学方法,检测VEGF和TP在大肠癌原发灶和相应淋巴结转移灶中的表达,共观察33例.结果 大肠癌原发灶中癌细胞TP及VEGF的阳性表达率与相应淋巴结转移灶中的表达无显著差异;7例预后较好的病例中,原发灶中间质细胞TP表达明显,淋巴结转移灶中癌细胞VEGF不表达.结论 大肠癌原发灶和相应淋巴结转移灶之间癌细胞的TP、VEGF表达水平无显著差异;对于有淋巴结转移的大肠癌,癌细胞TP表达水平不能预测而间质细胞及炎症细胞的TP表达水平可能预测癌对以5-FU为基础的化疗反应.

目的 探讨乳腺非典型性导管增生(ADH)与乳腺癌的关系.方法 收集乳腺癌和非乳腺癌标本共442例,其中浸润性癌253例,非浸润性癌(导管内原位癌)79例(低级别25例,高级别54例),非乳腺癌110例.观察ADH的形态及免疫组织化学染色情况;对各类病变的ADH检出率进行统计学分析.结果 ADH直径均小于2 mm;腺腔细胞CK5/6表达阴性,肌上皮细胞p63表达阳性.非浸润性癌ADH检出率显著高于非乳腺癌和浸润性癌(均P=0.000);非浸润性癌低级别病变的ADH检出率显著高于浸润性癌和非浸润性癌高级别病变(均P=0.000).结论 乳腺的ADH病变与导管原位癌的关系密切,及时发现和切除这种病变对预防乳腺癌发生有重要意义.

脂肪平滑肌瘤（包括子宫脂肪瘤）是一种少见、容易辨认、由平滑肌和成熟脂肪细胞密切混合而成的良性肿瘤，其包含一系列含有脂肪成分的病变，范围由含有少量脂肪成分到完全由脂肪细胞组成的肿瘤［1］。报道如下。

PURPOSE:To evaluate elastography using a bi-plane transducer for localizing prostate cancer (PCa) in patients scheduled for radical prostatectomy (RP), in comparison with step section pathological analysis.METHODS:Fifty-six consecutive PCa patients underwent real-time elastography examination with a bi-plane transducer before RP. Transverse elastographic images were obtained from the apex to the base by slightly compressing and releasing the prostate tissue using the probe. The diagnostic performance of elastography was evaluated in correlation with step section RP histopathology.RESULTS:In 56 PCa patients, gray-scale ultrasonography detected at least one lesion in 36 patients, whereas elastography detected at least one lesion in 53 patients (P = 0.001). The overall sensitivity, specificity and accuracy of elastography in depicting tumor lesions were 67.6, 89.5 and 82.7 %, respectively. The detection rate of a PCa lesion with elastography was best in the left posterior region, followed by the right posterior region. Elastography was more sensitive in detecting PCa lesions with higher Gleason scores, diameter >5 mm and extracapsular extension.CONCLUSIONS:The additional use of elastography with the bi-plane transducer can improve PCa detection rate by providing more information about tissue stiffness within the prostate gland.

目的 探讨儿童横纹肌肉瘤的病理形态学特点.方法 复习和分析30例儿童横纹肌肉瘤的形态学和免疫组织化学染色结果.结果 30例横纹肌肉瘤患儿中,男性22例,女性8例.组织学类型包括胚胎性(20/30,66.67％)、腺泡状(8/30,26.67％)和梭形/硬化性(2/30,6.67％)横纹肌肉瘤3种.发生部位依次为躯干(15/30,50.00％)、头颈(7/30,23.33％)、四肢(6/30,20.00％)和泌尿生殖系统(2/30,6.67％).免疫组织化学染色示肌特异性肌动蛋白(MSA)、平滑肌肌动蛋白(SMA)、结蛋白(DES)、肌细胞生成素(MyoG)和肌调节蛋白(MyoD1)均有不同程度的阳性表达.结论 胚胎性、腺泡状和梭形/硬化性横纹肌肉瘤是儿童横纹肌肉瘤的主要类型,组织学上主要表现为原始的小圆细胞,部分病例可见横纹肌母细胞.肿瘤细胞不同程度表达MSA 、SMA、DES、MyoG和MyoD1等肌源性标志物.鉴别诊断主要包括神经母细胞瘤、外周原始神经外胚层肿瘤等.

Objective To observe the morphological changes and significance of apocrine metaplasia with atypia in breast lesions.Methods The pathological sections of 190 cases of invasive breast cancer and 48 cases of breast benign lesions were retrospectively analysed between January 2011 and September 2012,the morphological characteristics of apocrine metaplasia were observed under microscope,and gross cystic disease fluid protein-15 kDa(GCDFP-15) and androgen receptor(AR) were determined by immunohistochemical staining.Results Apocrine metaplasia exhibited cystic,papillary and atypical type under microscope.There were 40 cases of apocrine metaplasia in invasive cancer,including 28cases of cystic type,4 cases of papillary type and 8 cases of atypical type.There were 13 cases of apocrine metaplasia in breast benign lesions,including 9 cases of cystic type,2 cases of papillary type and 2 cases of atypical type.There was no significant difference in the incidence of apocrine metaplasia between cases of invasive cancer and those of breast benign lesions(P>0.05).Apocrine metaplasia of atypical type accounted for 4.2%(8/190) and 4.2%(2/48) in cases of invasive cancer and those of breast benign lesions respectively,and accounted for 20.0%(8/40) and 15.4%(2/13) in cases of invasive cancer with apocrine metaplasia and those of breast benign lesions with apocrine metaplasia respectively.Immunohistochemical staining revealed that the expression of GCDFP-15 and AR was positive in apocrine metaplasia.Conclusion Apocrine metaplasia extensively exists in the benign and malignant lesions of the breast,and can be identified by expression of GCDFP-15 and AR.Though no significant difference is found between invasive breast cancer and breast benign lesions,apocrine metaplasia of atypical type has the risk to develop into breast cancer,which should be attached great importance.

Objective To determine whether cadence contrast pulse sequencing (CPS) harmonic ultrasonography can be used to predict aggressiveness of peripheral zone prostate cancer.Methods CPS harmonic ultrasonography was performed in 62 biopsy-proved prostate cancer patients.Time intensity curves were reconstructed for biopsy-proved or radical prostatectomy histopathology proved prostate cancer lesions.The characteristics of the curves were described using hemodynamic parameters including arrival time (AT),time-to peak (TTP) and peak intensity (PI).The differences of hemodynamic parameters between different Gleason score and pathologic stage were analyzed.Results Prostate biopsy revealed 156 peripheral zone prostate cancer lesions among 62 patients,including 40 low grade lesions and 116 high grade lesions.In comparison with low-grade lesions,the contrast agents arrived and distributed earlier in highgrade lesions (P =0.005,0.023).In addition to lower AT and TTP,high-grade tumors had higher PI than low-grade tumors (P =0.008).Among 21 patients underwent radical prostatectomy,histopathology documented 14 low grade tumors and 17 high grade tumors,significant differences of hemodynamic parameters were found between these two groups (P ＜0.05).Furthermore,prostate cancer lesions with extra capsular extension also had high PI than those confined in the gland (P =0.000).Conclusions Contrast-enhanced ultrasound and hemodynamic parameters might be helpful in predicting aggressiveness of prostate cancer.

目的 了解儿童卵黄囊瘤(YST)的组织形态学特征,明确免疫组织化学特点及其在YST病理学诊断中的作用,探讨YST鉴别诊断要点.方法 回顾总结21例YST(性腺8例,非性腺13例)的组织形态学及免疫组织化学特征.结果 YST的组织形态学表现多样,具有微囊和网状结构17例,内胚窦结构16例,腺管-腺泡结构10例,乳头状结构8例,巨囊结构和肠型结构各3例,多囊泡卵黄囊样结构2例,黏液瘤样结构1例.免疫组织化学染色甲胎蛋白(AFP)阳性表达20例(95.24％),细胞角蛋白(CK)阳性表达21例(100％),波形蛋白(Vim)阳性表达16例(76.19％).结论 儿童YST可发生于性腺(睾丸和卵巢),亦可发生于非性腺部位(骶尾部、盆腹腔、小脑和软组织等).YST组织形态表现具多样性,常见特征性结构有微囊和网状结构、内胚窦结构、腺管-腺泡结构和乳头状结构,巨囊结构、肠型结构、黏液瘤样结构和多囊泡卵黄囊样结构等较少见.组织形态学及免疫组织化学特征有助于明确YST的病理学诊断和鉴别诊断.

ObjectivesTo determine the accuracy of contrast-enhanced transrectal ultrasonography for tumor size measurements of hypoechoic prostate cancer foci located in the peripheral zone. MethodsA total of 55 men scheduled for radical prostatectomy, with biopsy-proven cancer in hypoechoic foci located in the peripheral zone, were consecutively enrolled in the present prospective study. Each patient underwent grayscale ultrasound and contrast-enhanced transrectal ultrasonography of the prostate according to a standardized protocol. The maximum tumor diameter on grayscale imaging and contrast-enhanced transrectal ultrasonography was compared with that determined using histopathology. ResultsA mean underestimation was documented to be approximately 3.9mm and 0.6mm for grayscale and contrast-enhanced transrectal ultrasonography imaging, respectively. Grayscale and contrast-enhanced transrectal ultrasonography imaging underestimated measurements by 76.67% (46 of 60) and 48.33% (29 of 60), whereas overestimated measurements were 20% (12 of 60) and 26.67% (16 of 60), respectively. A strong correlation was observed between contrast-enhanced transrectal ultrasonography and histopathological measurements (r=0.91, P<0.0001). A weak linear correlation was found between grayscale and histopathological measurements (r=0.59, P<0.0001). Bland-Altman analysis results were in complete accordance with correlation analysis results. For cases with maximum histopathological tumor diameters 10mm and >10mm, 40% (6 of 15) and 86.67% (39 of 45) were index tumors, respectively (P<0.0001). ConclusionsContrast-enhanced transrectal ultrasonography is significantly more accurate than conventional grayscale imaging for measuring prostate tumor size, especially for tumors with a diameter >10mm, and it might have a role in preoperative assessment of prostatic index tumor sizes.

Breast atypical ductal hyperplasia(ADH) has the risk of progression into breast cancer,and there are characteristic molecular biological and genetic changes in ADH,which is to some extent similar to ductal carcinoma in situ(DCIS).The increase of cell proliferation index,hormone receptor expression and cell growth regulating factors may promote the formation of ADH.Chromosome abnormality and epigenetic changes may lead to the formation of ADH,among which 16p,17q chromosome abnormality and promoter methylation of cancer related gene may be the early events of ADH.Studies have confirmed that ADH is the precursor of low grade ductal carcinoma in situ(LG-DCIS) which shares the similar molecular biological characteristics and genetic performance with ADH,and can progress to low grade invasive ductal carcinoma(LG-IDC) through LG-DCIS stage.Studies have also pointed out that ADH is not the precursor of high grade ductal carcinoma in situ(HG-DCIS),and high grade invasive ductal carcinoma(HG-IDC) has different pathogenesis.With the abnormal expression of cell adhesion molecule,extracellular matrix produces a large number of matrix protease which results in reduced cell-to-cell adhesion and degradation of basement membrane,and ultimately promotes the development of ADH to IDC.

目的:探讨前列腺小细胞癌的临床病理特点、诊断及鉴别诊断。方法:收集5例原发性前列腺小细胞癌,对其临床特点、组织学形态及免疫组化进行分析,并复习相关文献。结果:5例患者年龄52岁-81岁,平均年龄69.8岁,中位年龄73岁。主诉均为排尿困难。1例有前列腺腺癌内分泌治疗病史。血清总PSA正常或增高。组织学形态均表现为弥漫成片或巢状排列的卵圆形及短梭形细胞。2例合并前列腺腺癌。免疫组化,4例表达AE1/AE3,2例表达CK,3例表达TTF1,5例均表达Syn、CgA、CD56、NSE,5例均不表达PSA、PAP、AR、P504S、LCA、Des。讨论:前列腺小细胞癌是罕见的前列腺恶性肿瘤,需与多种小细胞恶性肿瘤鉴别。预后较差。

患儿男,5岁,因反复尿痛2年、加重2个月入院.病程中无血尿,偶有脐周疼痛.B超示膀胱右侧壁见实质性肿块,呈高回声,38 mm×29 mm大小,内部回声不均,见数个无回声区.膀胱活检:黏膜慢性炎,伴嗜酸性粒细胞浸润.予抗炎及口服激素治疗,肿块无明显缓解.遂行膀胱肿块切除术. 病理检查 带蒂分叶状肿块,4 cm×3 cm×2 cm大小,蒂直径0.3 cm,切面淡黄、灰红色,质嫩,部分半透明,切面见数个囊腔,直径0.5～1 cm,内为淡黄色清液.镜检:黏膜上皮呈腺性膀胱炎及囊性膀胱炎,见类似Von Brunn巢的上皮聚集(图1),局部伴有鳞化,黏膜内淋巴细胞、浆细胞及少量嗜酸性粒细胞浸润,部分黏膜呈纤维上皮性息肉(图2),间质为梭形及星芒状的纤维母细胞及排列紊乱的平滑肌纤维,未见核分裂.部分血管玻璃样变.免疫表型:SMA阳性,平滑肌desmin阳性,Ki-67、MyoG和MyoD1均阴性.

Background Rhabdomyosarcoma (RMS) is an uncommon malignancy of the breast. The aim of this study was to summarize its clinicopathologic features and biological behavior. Methods Five primary or secondary breast RMSs were collected. Their clinicopathological characteristics and all published literature about breast RMS were reviewed. Immunohistochemical study of desmin, myogenic differentiation 1 (MyoD1), myogenin, leukocyte common antigen (LCA), vimentin, cytokeratin (AE1/AE3), E-cadherin, neuron specific enolase (NSE), CD99, chorioallantoic membrane 5.2 (CAM5.2) and epithelial membrane antigen (EMA) expression were performed. Results The five patients were all female with ages ranging from 16 to 46 years old (mean, 30 years). Three were metastatic breast RMSs, two embryonal and one solid variant alveolar, with the primary tumor sites the right labium majus, left nasal meatus and nasopharynx, respectively. The other two, one embryonal and one alveolar, were primaries. Grossly, the surgical specimens revealed round or oval, well-demarcated but nonencapsulated masses. Their cut surfaces consisted of homogeneous grayish yellow or white tissue. Microscopically, most tumor cells were poorly differentiated small round, oval or small polygons with eosinophilic cytoplasm. All cases were positive for vimentin, desmin, MyoD1 and myogenin. One embryonal RMS also had a few cells with perinuclear staining of AE1/AE3. The other markers were negative. Conclusions Although primary or metastatic RMS in breast was almost confined to young adolescent females, our cases suggested that it can also happen to the middle-aged women. Embryonal RMS has a certain metastatic potential. MyoD1 and myogenin are two useful markers when making differential diagnosis. Axillary lymph node status and age may play a role in the prognosis of primary breast RMS patients. Chin Med J 2012;125(14):2618-2622

Objective To establish an animal model for hypothyroidism and thyroid-associated ophthalmopathy(TAO) by method of the human thyrotropin(hTSH) stimulus and radioactive iodine-131(131I) injection in Wistar rats.Methods Twenty Wistar rats were divided randomly into experimental group and normal group(group D),the experimental group were injected intraperitoneally with hTSH 8 weeks of intraperitoneal injection 131I,based on different dosage of 131I,experiments groups were divided into low-dose group(group A),medium dose group(Group B) and the high-dose group(Group C).All rats were sacrificed at weeks 16 for histology,thyroid and orbital tissue were isolated,observation and analysis of pathological changes in rat orbit with routine HE staining.Serum FT3and FT4levels were detected by electro chemiluminescence,serum rat TSH(rTSH) and TRAb levels measured by ELISA.Results After eight weeks of injection 131I,serum FT4 level in group A(16.98±2.92) pmol/L,group B(1.84± 0.44) pmol/L and group C(1.35± 0.37) pmol/L are reduced,three groups and normal group(group D)differences were significant(P<0.05);group C and group B were significantly lower than group A,the difference was significant(P<0.05);serum rTSH levels in group C(2.53±0.10)mIU/L and group B(2.37±0.32) mIU/L were slightly higher as compared to group D(2.17±0.38) mIU/L,but the differences were significant(P>0.05);group A(1.78±0.36) mIU/L is slightly lower than group D,but also no significant difference(P>0.05).Serum TRAb level in group A(4.69±1.52)IU/ml and group B(4.49±2.60)IU/ml was significantly lower than that in group D(12.79±0.75)IU/ml,the difference was significant(P>0.05).group C(10.36±4.48)IU/ml and group D was no significant difference(P>0.05).Histological observation in orbital tissues,experimental groups displayed edema and mucinous degeneration,intercellular space widening significantly,mast cell infiltration in most of rats.With increasing doses of 131I,group B and group C showed proliferation of adipose tissue and fibrous tissue,degeneration and disruption of muscular fibers,muscle fiber fracture,orbital vascular dilatation,the muscle cell nucleus hypertrophy.The vacuole formation appeared in muscle fibers were found in group C.In group D,orbital tissues were normal histologically.Conclusion The animal model constructed by hTSH stimulus and131I injection produced general characteristics of hypothyroidism and TAO.And confirmed the presence of hypothyroidism associated with TAO.The method is simple and to utilize not only for fundamental research of hypothyroidism after 131I treatment,but also as a model to explore other reasons caused hypothyroidism or the TAO mechanism.

Objective To investigate the utility of the transition zone index (TZI) for optimal predictor selection of prostate cancer (PCa) in patients with prostate-specific antigen (PSA) levels of 4 to 10 ng/ml. Methods In this retrospective cohort study, results of transrectal ultrasonography (TRUS)-guided biopsy were assessed in 616 consecutive patients; The prostate and transition zone volumes were determined by TRUS. A TZI cutoff value of 0.47 produced the best sensibility and specificity rates in receiver operating characteristic (ROC) curve analysis and thus was used to classify the study subjects into group TZI ≤0.47 and group TZI ≤0.47. Logistic regression analysis was used to predict outcomes. The variables that were statistically significant in the stepwise logistic regression analysis were assessed using the ROC curve and the area under the curve (AUC). Results Overall, 166 of the 616 patients (26.9%) had histologically confirmed PCa. A total of 238 (38.6%) patients were classified into group TZI ≤0.47, of whom 97 (40.8%) exhibited a positive biopsy, and 378 (61.4%) patients were classified into group TZI>0.47, of whom 69 (18.3%) exhibited a positive biopsy. The stepwise logistic regression analysis revealed that PSA density (PSAD) exhibited the strongest predictive value in the overall population and in group TZI ≤0.47, whereas PSA transition zone density (PSATZD) was the optimal predictor in group TZI>0.47. The ROC curve analysis revealed that when using the TZI- specific 100% sensitivity cutoffs, 17.7% and 25% of the biopsies were unnecessary and could be avoided in the overall patient population prior to and following the division into groups, respectively (P = 0.002). Using an individually generated 95% sensitivity cutoff of 0.12 ng/ml 2 for PSAD and a cutoff of 0.179 ng/ml 2 for PSATZD for TZI-stratified cohorts of TZI ≤ 0.47 and TZI>0.47, a more consistent specificity of 44% and 46.9%, respectively, for each cohort was observed. Conclusion The optimal predictor for PCa differs between various TZI levels. The combination of PSAD in patients with TZI≤0.47 and PSATZD in patients with TZI>0.47 helps to identify potentially unnecessary biopsies compared to the use of a single PSAD for the entire patient population.

Objective To determine the utility of routine transition zone biopsies in men undergoing transrectal ultrasound-guided transperineal prostate biopsies. Methods A total of 1956 consecutive patients underwent transrectal ultrasound-guided prostate biopsies for the first time. Systematic biopsies and additional 2 cores transition zone biopsies were performed. The patients were classified to different subgroups according to prostate-specific antigen (PSA), digital rectal examination (DRE) results, transrectal ultrasonography (TRUS) results, prostate volume and PSA density (PSAD). The impact of transition zone biopsies on the prostate cancer detection rate of different groups of patients was analyzed. Results Prostate cancer was detected in 762(39.0%)of the 1956 patients, and 86 of these patients had positive cores only in TZ (P=0.004). The increase in the cancer detection rate by two-core TZ biopsies was 5.8%, 5.5%, 6.8%, 4.3%and 5.8% respectively in patients with PSA level above 10 ng/ml, negative DRE findings, negative TRUS findings, prostate volume over 30ml and PSAD above 0.18 (P<0.05). Conclusion Routine transition zone biopsies may improve the detection rate of prostate cancer in transperineal biopsy protocol.

目的探讨移行区指数(TZI)在前列腺特异性抗原(PSA)灰区(4~10ng/ml)前列腺癌(PCa)预测参数优化选择中的作用.方法对616例PSA位于灰区并接受经直肠超声(TRUS)引导的前列腺穿刺活检患者的临床资料进行回顾性分析.TRUS测量前列腺体积(PV)及其移行区体积(TZV).运用TZI在ROC曲线中诊断敏感性及特异性最佳的界值0.47将总研究样本分为TZI≤0.47组和TZI>0.47组.Logistic回归分析确定各组中的PCa最佳预测参数,并对各组独立预测参数的ROC曲线及其曲线下面积(AUC)进行分析.结果616例患者中,穿刺病理诊断166例(26.9%)为PCa,其余450例(73.1%)为良性病变.TZI≤0.47组包括238例(38.6%)患者,其中97例(40.8%)患者为PCa;TZI>0.47组包括378例(61.4%)患者,其中69例(18.3%)患者为PCa.对TZI≤0.47的患者,前列腺特异性抗原密度(PSAD)为最佳预测参数,其95%敏感性诊断界值为0.12 ng/ml 2;而对TZI>0.47的患者,移行区前列腺特异性抗原密度(PSATZD)为最佳预测参数,其95%敏感性诊断界值为0.179 ng/ml 2.最佳预测参数取100%敏感性时的诊断界值,TZI分组前后可避免的不必要的前列腺穿刺分别为17.7%及25%(P=0.002).结论不同TZI组别中PCa最佳预测参数不同,对TZI≤0.47患者使用PSAD,而对TZI>0.47的患者使用PSATZD分别作为最佳预测参数较分组前仅使用PSAD作为最佳预测参数能使更多的患者避免不必要的穿刺活检.

Objective To investigate the clinical significance of pyrosequencing assay for determining K-ras mutations in exon 2 codons 12 and 13 in clinical colorectal cancer tissues.Methods Genomic DNA,extracted from K-ras mutant cell lines SW480 (homozygous,c.35G ＞ T), DLD-1 (heterozygous,c.38G ＞ A) and wild-type HT-29,was first used as the sequencing template respectively to test the accuracy of pyrosequencing methodology.The SW480 and DLD-1 DNA was separately mixed with wild-type HT-29 DNA in proportions of 2％,3％,5％,10％,20％,30％ and 50％,the sensitivity for mutation detection was measured separately by pyrosequencing assay and directed Sanger DNA sequencing in the serial DNA mixture samples.The pyrosequencing assay results were compared with the corresponding Sanger sequencing and the datas were analysized by Fisher exact test.Pyrosequencing analysis was then performed for screening K-ras exon 2 mutations at codons 12 and 13 on DNA isolated from a panel of 30 colorectal cancer samples derived from clinical formalin-fixed and paraffin embedded (FFPE)tissues.Results Cancer cell lines with known K-ras mutations ( SW480 and DLD-1 ) were readily detectable by pyrosequencing-based analysis.When the proportions of mutant colorectal cancer cell line DNA were 5％ and 10％ content,the mutation rates of K-ras gene detected by conventional Sanger DNA sequencing were 33.3％ (4/12) and 58.3％ (7/12) respectively,whereas the mutation rates detected by pyrosequencingbased assay were 91.7％ (11/12) and 100％ (12/12) respectively,there were significant differences between those two sequencing methodology ( P ＜0.05).Furthermore,we found 10 patients with K-ras exon 2 point mutations at codons 12 and 13 by pyrosequencing-based assay from 30 colorectal cancer FFPE tissues,the point mutation rate was 33.3％ (10/30) and all of the mutations determined were heterozygous.The codon 12 was most frequently affected [30％ (9/30)].Mutations with the highest frequency were G ＞ A transitions [ 50％ ( 5/10 ) ],followed by G ＞ T transversions [ 30％ ( 3/10 ) ].Conclusion The pyrosequencing assay provides an accurate and sensitive method for mutation screening of K-ras exon 2 codons 12 and 13 in routine diagnostic specimens,thereby allowing the selection of the cancer treatment in clinical individualized practice.