Fusion expression, purification, and characterization of cytokeratin 19 fragments in E. coli for enhanced stability in diagnostic applications.

Cytokeratin 19 fragment (CYFRA21-1) serves as a crucial tumor marker in the context of lung cancer patients, playing a pivotal role as a calibrator in the realm of in vitro diagnostics. Nevertheless, during practical application, it has come to light that the recombinantly synthesized full-length CYFRA21-1 antigen exhibits suboptimal stability at the requisite concentration, while the utilization of natural antigens incurs a substantial cost. To address this issue, our investigation harnessed a strategic approach whereby the soluble fragment of cytokeratin 19 (Aa244-400) was integrated into the pET32a vector, subsequently being expressed within E. coli through a fusion with the TrxA protein. This process involved induction of protein expression through 0.2 mM IPTG at 16 °C for a duration of 16 h. After induction, the target protein was purified through Ni affinity and ion exchange chromatography. Subsequent characterization of the targeted protein was executed through the SEC-HPLC technique. The attained CYFRA21-1 antigen, as generated within this study, was effectively incorporated into a chemiluminescence-based in vitro diagnostic detection kit. The results indicate that the fusion protein exhibited commendable reactivity and stability, manifesting a deviation of less than 10 % following incubation at 37 °C for 7 days. Importantly, the production yield achieved a notable magnitude of 300 mg/L, thus rendering it a cost-effective and scalable alternative to natural antigens for clinical diagnostic applications.

Evaluation of the follicular patterned thyroid lesions based on the WHO 2022 criteria with an emphasis on the grey-zone lesions.

Follicular-patterned thyroid nodules (FPTN) are classified byWHO-2022 into benign, borderline and malignant categories. There are however, grey-zone lesions that pose a diagnostic challenge due to ambiguity in defining criteria and inter-observer variability. WHO-2022 has enumerated specific diagnostic criteria for these lesions. Accurate categorization of morphologically similar TNs is vital to reduce overtreatment of indolent lesions. In this study, we have reclassified FPTNs according to WHO-2022 criteria, emphasizing on grey-zone lesions. We studied the utility of immunohistochemistry (IHC)-CD56, HBME-1 and CK19 in distinguishing benign from malignant nodules and BRAFV600E IHC to better distinguish the (widely-invasive) encapsulated follicular variant of papillary thyroid carcinoma (FVPTC) from infiltrative FVPTC. Only those cases with dominant nodule having follicular pattern histology were included and re-evaluated for following histopathological features-focality, encapsulation, circumscription, nuclear PTC features, capsular-invasion, angio-invasion, papillae and necrosis. IHC findings for above-mentioned markers were noted. Seventy-nine cases met the inclusion criteria. Amendment of original diagnosis was done in 19 % cases. BRAFV600E IHC was positive in the two cases of infiltrative FVPTC while it was negative in all nine IE (invasive encapsulated) FVPTCs. Diffuse HBME1 was noted in most malignant nodules (61 %) while CD56 was expressed more often in benign lesions (70 %). CK19 was positive in lesions displaying nuclear PTC features (86 %). Using WHO 2022 criteria, we were able to re-classify follicular thyroid lesions with greater confidence. Appropriate IHC panel in adjunct to histology aids in categorizing challenging cases.

Odontogenic carcinoma with dentinoid: case report and literature review of a rare entity.

BACKGROUND: Odontogenic carcinoma with dentinoid (OCD) is a rare and controversial entity, which has not yet been included in the current World Health Organization classification of odontogenic lesions. Owing to the small number of reported cases, the clinicopathological characteristics, biological behavior, prognosis, and appropriate treatment strategies for OCD remain to be defined. Herein, we present an additional case of OCD with a focus on the differential diagnosis and review of the pertinent literature, in order to enable better recognition by oral clinicians and pathologists and further characterization of this entity. CASE PRESENTATION: This paper reports a case of OCD in the posterior mandible of a 22-year-old female. Radiography showed a well-defined unilocular radiolucency with radiopaque materials. The intraoperative frozen section pathology gave a non-committed diagnosis of odontogenic neoplasm with uncertain malignant potential. Then a partial mandibulectomy with free iliac crest bone graft and titanium implants was performed. Microscopically, the tumor consisted of sheets, islands, and cords of round to polygonal epithelial cells associated with an abundant dentinoid matrix. Immunohistochemically, the tumor cells were diffusely positive for CK19, p63, and ß-catenin (cytoplasmic and nuclear). No rearrangement of the EWSR1 gene was detected. The final diagnosis was OCD. There has been no evidence of recurrence or metastasis for 58 months after surgery. We also provide a literature review of OCD cases, including one case previously reported as ghost cell odontogenic carcinoma from our hospital. CONCLUSIONS: OCD is a locally aggressive low grade malignancy without apparent metastatic potential. Wide surgical excision with clear margins and long-term period follow-up to identify any possible recurrence or metastases are recommended. Histopathological examination is essential to conclude the diagnosis. Special care must be taken to distinguish OCD from ghost cell odontogenic carcinoma and clear cell odontogenic carcinoma, as misdiagnosis might lead to unnecessary overtreatment. Study of additional cases is required to further characterize the clinicopathological features and clarify the nosologic status and biological behavior of this tumor.

The value of MRI in predicting hepatocellular carcinoma with cytokeratin 19 expression: a systematic review and meta-analysis.

AIM: To evaluate the overall diagnostic performance of magnetic resonance imaging (MRI), different image features, and different image analysis methods in predicting hepatocellular carcinoma (HCC) with cytokeratin 19 (CK19) expression. MATERIALS AND METHODS: A systematic literature search was performed to identify studies using MRI to predict HCC with CK19 expression between 2012 and 2023. Data were extracted to calculate the pooled sensitivity and specificity. Overall diagnostic performance was assessed using areas under the summary receiver operating characteristic curve (AUC). Subgroup analyses were conducted for specific image features and according to image analysis methods (traditional image feature, radiomics, and combined methods). Z-test statistics was used to analyse the differences in diagnostic performance between combined and individual methods. RESULTS: Eleven studies with 14 datasets (1,278 lesions from 1,264 patients) were included. The overall pooled sensitivity, specificity, and AUC with corresponding 95% confidence intervals were estimated to be 0.72 (0.55, 0.85), 0.88 (0.80, 0.93), and 0.89 (0.86, 0.91) for MRI in predicting HCC with CK19 expression. Combined methods had higher sensitivity than image feature methods (0.86 versus 0.54, p=0.001), with no difference in specificity (0.85 versus 0.87, p=0.641). There were no significant differences between radiomics and combined methods regarding sensitivity (p=0.796) and specificity (p=0.535), respectively. CONCLUSION: MRI shows moderate sensitivity and high specificity in identifying HCC with CK19 expression. The application of radiomics can improve the sensitivity of MRI in identifying HCC with CK19 expression.

Regional Differences in Human Biliary Tissues and Corresponding In Vitro-Derived Organoids.

BACKGROUND AND AIMS: Organoids provide a powerful system to study epithelia in vitro. Recently, this approach was applied successfully to the biliary tree, a series of ductular tissues responsible for the drainage of bile and pancreatic secretions. More precisely, organoids have been derived from ductal tissue located outside (extrahepatic bile ducts; EHBDs) or inside the liver (intrahepatic bile ducts; IHBDs). These organoids share many characteristics, including expression of cholangiocyte markers such as keratin (KRT) 19. However, the relationship between these organoids and their tissues of origin, and to each other, is largely unknown. APPROACH AND RESULTS: Organoids were derived from human gallbladder, common bile duct, pancreatic duct, and IHBDs using culture conditions promoting WNT signaling. The resulting IHBD and EHBD organoids expressed stem/progenitor markers leucine-rich repeat-containing G-protein-coupled receptor 5/prominin 1 and ductal markers KRT19/KRT7. However, RNA sequencing revealed that organoids conserve only a limited number of regional-specific markers corresponding to their location of origin. Of particular interest, down-regulation of biliary markers and up-regulation of cell-cycle genes were observed in organoids. IHBD and EHBD organoids diverged in their response to WNT signaling, and only IHBDs were able to express a low level of hepatocyte markers under differentiation conditions. CONCLUSIONS: Taken together, our results demonstrate that differences exist not only between extrahepatic biliary organoids and their tissue of origin, but also between IHBD and EHBD organoids. This information may help to understand the tissue specificity of cholangiopathies and also to identify targets for therapeutic development.

Electrochemical ultrasensitive detection of CYFRA21-1 using Ti3C2Tx-MXene as enhancer and covalent organic frameworks as labels.

The concentration level of cytokeratin fragment antigen 21-1 (CYFRA21-1) can be used as an important indicator for predicting non-small cell lung cancer (NSCLC). Here, a sandwich-type electrochemical immunosensor for ultrasensitive detection of CYFRA21-1 is developed. The sensor based on a combination of gold nanoparticle (AuNPs) decorated Ti3C2Tx-MXene (Au-Ti3C2Tx) as the substrate enhancer, and toluidine blue (TB) modified AuNPs doped covalent organic framework (COF) polymer as the signal tag (TB-Au-COF). The Au-Ti3C2Tx is used to capture numerous primary antibodies and accelerate the electron transfer rate of the substrate, while the TB-Au-COF can be applied to provide a large number of signal units TB and secondary antibodies. These features of composites endow the proposed immunosensor with high sensitivity and current response to CYFRA21-1. Under optimum conditions, the immunosensor offers a wide current response for CYFRA21-1 from 0.5-1.0 × 104 pg·mL-1 with a detection limit of 0.1 pg·mL-1. Furthermore, the biosensing platform can be applied for CYFRA21-1 detection to analyze real serum samples, providing an effective and useful avenue for the applicability of Au-Ti3C2Tx and TB-Au-COF composite materials in biosensing field.

A rare case of high-grade intraductal carcinoma of the upper lip: immunohistochemical and genetic analyses.

Although intraductal carcinoma (IDC) of the salivary glands was previously called low-grade cribriform cystadenocarcinoma, it was newly categorized in the 4th version of the World Health Organization classification. We report a case of IDC of the upper lip and examined it immunohistochemically and genetically. The patient was a 48-year-old Japanese female, who noticed a tiny nodule on her left upper lip. Histologically, the tumor cells, which had eosinophilic cytoplasm, exhibited papillary and solid growth patterns, and regions of suspected microinvasion or intraductal spread were also seen at the periphery of the tumor. Small necrotic foci were noted. Immunohistochemically, the tumor cells were diffusely positive for the androgen receptor, CK19, CK5/6, EGFR, and SOX10, whereas they were focally positive for GCDFP-15, S-100 protein, and mammaglobin. The tumor nests were surrounded by alpha-smooth muscle actin-p63-/calponin-/CK14-positive myoepithelial cells. The Ki-67 labeling index was 51.2%. Genetic analysis showed no evidence of the TRIM27-RET or NCOA4-RET fusion gene. We finally diagnosed the tumor as a high-grade mixed intercalated duct/apocrine-type IDC of the upper lip. IDC of the minor salivary glands is exceedingly rare. We discuss diagnostic problems associated with minor salivary gland lesions, and the "basal-like" phenotype of this case.

Self-Supply of H2O2 and O2 by Hydrolyzing CaO2 to Enhance the Electrochemiluminescence of Luminol Based on a Closed Bipolar Electrode.

A novel luminol electrochemiluminescence (ECL) sensing platform was developed based on the closed indium tin oxide bipolar electrode (ITO BPE) for detecting the biomarkers of squamous cell carcinomas named cytokeratin 19 fragments (CYFRA 21-1). A strategy of in situ-generated coreactant H2O2 and O2 was proposed to enhance luminol ECL intensity. CaO2, possessing a high capacity of self-supplying H2O2 and O2, was encapsulated by ZIF-67, glucose oxidase, and horseradish peroxidase (GOD/HRP-loaded ZIF-67@CaO2). In the presence of glucose, gluconic acid and H2O2 were generated via the catalytic effect of GOD; then, gluconic acid induced the degradation of ZIF-67, leading to the hydrolysis of the unprotected CaO2 to produce both O2 and H2O2. Therefore, the generated coreactants O2 and H2O2, via the oxidation of glucose and the hydrolysis of CaO2, can effectively enhance the luminol ECL behavior. According to the luminol ECL reaction occurring on the anode surface of the ITO BPE, a sandwich immunosensor was fabricated on the anodic ITO BPE and GOD/HRP-loaded ZIF-67@CaO2 as labels incubated with secondary antibodies. Owing to the cathodic reaction rate influencing the anodic reaction rate, Au nanoparticles were electrodeposited on the cathode surface of the ITO BPE, which not only meliorated oxygen reduction but also further enhanced the ECL intensity of luminol. Under optimal experiments, an ECL immunosensor for detecting CYFRA 21-1 with a wider linear range from 0.0075 to 50 ng mL-1 and a detection limit of 1.89 pg mL-1 (S/N = 3) was obtained.

Etching Triangular Silver Nanoparticles by Self-generated Hydrogen Peroxide to Initiate the Response of an Electrochemiluminescence Sensing Platform.

This work outlines a versatile and high-performance electrochemiluminescence (ECL) platform that uses complex luminescent molecules [Ru(II) complex] formed by carbohydrazide (CON4H6) and tris(4,4'-dicarboxylicacid-2,2'-bipyridyl)ruthenium(II) dichloride [Ru(dcbpy)32+] as emitters to facilitate the intramolecular ECL mechanism for reducing the response distance and interference, and they were kept immobilized on a porous bismuth vanadate nanoarray (BiVO4 NA) to improve the orderliness of electron transfer. In addition, the detection was made depending on the etching of triangular silver nanoparticles (T-Ag NPs) by self-generated hydrogen peroxide (H2O2) to initiate the recovery response of the originally quenched ECL due to ECL-RET between the Ru(II) complex (donor) and T-Ag NPs (receptor). Because of the antibacterial application of dopamine, its own redox ability could produce more H2O2 for etching receptor T-Ag NPs under near-infrared (NIR) stimulation. Notably, in this system, the specific binding of antigens and antibodies with the autogenesis process of H2O2 and the ECL detection procedure are independent. Therefore, the proposed system can avert the impact of complex biological samples effectively, and the ECL efficiency of the Ru(II) complex can be readily utilized. On this basis, a biosensor is explored for the primary diagnosis of squamous cell carcinoma by detecting the biomarker named after cytokeratin fragment 19 (CYFRA21-1), from which an excellent linearity from 0.1 pg/mL to 50 ng/mL is achieved with a detection limit of 0.058 pg/mL. All of these results confirmed that this strategy can be a promising candidate for fabricating an ECL-based biosensor.

Intramolecular Coreaction Accelerated Electrochemiluminescence of Polypeptide-Biomineralized Gold Nanoclusters for Targeted Detection of Biomarkers.

The development of label-free electrochemiluminescence (ECL)-based sensing technology for biomarkers detection has a congenital defect compared to noncompetitive sandwich-type biosensors due to the lack of detection antibody conjugated with a signal label. Nevertheless, it is still not difficult to realize the ultrasensitive analysis benefit from the exploration of efficient sensing substrates and signal transducers. In this work, an innovative sensing system is purposed utilizing Fe2O3 nanoarrays (Fe2O3 NAs) as a well-ordered coreaction accelerator and polypeptide-biomineralized gold nanoclusters (Au NCs) as a signal transducer. Bifunctional peptide ligands of H2N-MMYYHFRRHL-COOH (MYH-10) are self-designed; it cannot only play a role of reductant and coupling reagent for cluster formation using the MMYY sequence root in the N-terminal but also act as a connection for coupling carriers and immune molecules via the HFRRHL region of the C-terminal. In addition to intramolecular ECL emission between Au NCs and tris(3-aminoethyl)amine (TAEA), all strategies undoubtedly reduce the spatial hindrance of the sensing interface and increase the effectiveness of the electron transfer and immune recognition. With CYFRA21-1 as a target, the biosensor exhibits a linear ECL response in a wide range (10 fg mL-1 to ∼100 ng mL-1) and an ultralow detection limit of 1.33 fg mL-1 (S/N = 3). With convincing experimental data, these innovative strategies will be more eye-catching in peptide-based nanocluster synthesis and expansion of a more novel thought for sensing platform fabrication.

Three-dimensional visualization of cleared human pancreas cancer reveals that sustained epithelial-to-mesenchymal transition is not required for venous invasion.

Venous invasion is three times more common in pancreatic cancer than it is in other major cancers of the gastrointestinal tract, and venous invasion may explain why pancreatic cancer is so deadly. To characterize the patterns of venous invasion in pancreatic cancer, 52 thick slabs (up to 5 mm) of tissue were harvested from 52 surgically resected human ductal adenocarcinomas, cleared with a modified iDISCO method, and labeled with fluorescent-conjugated antibodies to cytokeratin 19, desmin, CD31, p53 and/or e-cadherin. Labeled three-dimensional (3D) pancreas cancer tissues were visualized with confocal laser scanning or light sheet microscopy. Multiple foci of venous and even arterial invasion were visualized. Venous invasion was detected more often in 3D (88%, 30/34 cases) than in conventional 2D slide evaluation (75%, 25/34 cases, P < 0.001). 3D visualization revealed pancreatic cancer cells crossing the walls of veins at multiple points, often at points where preexisting capillary structures bridge the blood vessels. The neoplastic cells often retained a ductal morphology (cohesive cells forming tubes) as they progressed from a stromal to intravenous location. Although immunolabeling with antibodies to e-cadherin revealed focal loss of expression at the leading edges of the cancers, the neoplastic cells within veins expressed e-cadherin and formed well-oriented glands. We conclude that venous invasion is almost universal in pancreatic cancer, suggesting that even surgically resectable PDAC has access to the venous spaces and thus the ability to disseminate widely. Furthermore, we observe that sustained epithelial-mesenchymal transition is not required for venous invasion in pancreatic cancer.

Noninvasive prediction of HCC with progenitor phenotype based on gadoxetic acid-enhanced MRI.

OBJECTIVES: To explore the noninvasive prediction of hepatocellular carcinoma (HCC) with progenitor phenotype based on gadoxetic acid-enhanced magnetic resonance imaging (MRI). METHODS: This retrospective study included 115 surgery-proven HCCs with preoperative gadoxetic acid-enhanced MRI from August 2015 to September 2018. Image features were reviewed. Quantitative image analysis was performed using histogram analysis. HCC with progenitor phenotype was defined as positive for either cytokeratin 19 (CK19) or epithelial cell adhesion molecule (EpCAM) expression. Statistically significant variables for identifying HCCs with progenitor phenotype were determined at multivariate analyses. ROC analyses were used to determined cutoff values and the diagnostic performance of significant variables and combinations. Prediction nomogram was constructed based on multivariate analysis. RESULTS: At multivariate regression analyses, AFP ≥ 155.25 ng/mL (p < 0.001), skewness on T2WI ≤ 1.10 (p = 0.024), uniformity on pre-T1WI ≤ 0.91 (p = 0.024), irregular tumor margin (p = 0.006), targetoid appearance (p = 0.001), and the absence of mosaic architecture (p = 0.014) were significant predictors of HCCs expressing progenitor cell markers. Combing any three of those significant variables, it provides a diagnostic accuracy of 0.86 (95% CI 0.78-0.92) with sensitivity of 0.97 (95% CI 0.86-1.00), and specificity of 0.74 (95% CI 0.63-0.83). The C-index of the regression coefficient-based nomogram was 0.94 (95% CI 0.91-0.98). CONCLUSIONS: Noninvasive prediction of HCCs with progenitor phenotype can be achieved with high accuracy by integrated interpretation of biochemical and radiological information, representing a handy tool for precise patient management and the prediction of prognosis. KEY POINTS: • Qualitative image features of irregular tumor margin, targetoid appearance, and the absence of mosaic architecture are significant predictors of hepatocellular carcinoma with progenitor phenotype. • Quantitative analyses using whole-lesion histogram analysis provides additional information for the prediction of hepatocellular carcinoma with progenitor phenotype. • Noninvasive prediction of hepatocellular carcinoma with progenitor phenotype can be achieved with high accuracy by integrated interpretation of clinical information and qualitative and quantitative imaging analyses.

A radiomics-based biomarker for cytokeratin 19 status of hepatocellular carcinoma with gadoxetic acid-enhanced MRI.

OBJECTIVES: We aimed to develop a radiomics-based model derived from gadoxetic acid-enhanced MR images to preoperatively identify cytokeratin (CK) 19 status of hepatocellular carcinoma (HCC). METHODS: A cohort of 227 patients with single HCC was classified into a training set (n = 159) and a time-independent validated set (n = 68). A total of 647 radiomic features were extracted from multi-sequence MR images. The least absolute shrinkage and selection operator regression and decision tree methods were utilized for feature selection and radiomics signature construction. A multivariable logistic regression model incorporating clinico-radiological features and the fusion radiomics signature was built for prediction of CK19 status by evaluating area under curve (AUC). RESULTS: In the whole cohort, 57 patients were CK19 positive and 170 patients were CK19 negative. By combining 11 and 6 radiomic features extracted in arterial phase and hepatobiliary phase images, respectively, a fusion radiomics signature achieved AUCs of 0.951 and 0.822 in training and validation datasets. The final combined model integrated a-fetoprotein levels, arterial rim enhancement pattern, irregular tumor margin, and the fusion radiomics signature, with a sensitivity of 0.818 and specificity of 0.974 in the training cohort and that of 0.769 and 0.818 in the validated cohort. The nomogram based on the combined model showed satisfactory prediction performance in training (C-index 0.959) and validation (C-index 0.846) dataset. CONCLUSIONS: The combined model based on a fusion radiomics signature derived from arterial and hepatobiliary phase images of gadoxetic acid-enhanced MRI can be a reliable biomarker for CK19 status of HCC. KEY POINTS: • Arterial rim enhancement pattern and irregular tumor margin on hepatobiliary phase on gadoxetic acid-enhanced MRI can be useful for evaluating CK19 status of HCC. • A radiomics-based model performed better than the clinico-radiological model both in training and validation datasets for predicting CK19 status of HCC. • The nomogram based on the fusion radiomics signature can be easily used for CK19 stratification of HCC.

Standard ultra-staging compared to one-step nucleic acid amplification for the detection of sentinel lymph node metastasis in endometrial cancer patients: a retrospective cohort comparison.

INTRODUCTION: The objective of this study was to compared standard ultra-staging (SU) with one-step nucleic acid amplification (OSNA) for the detection of sentinel lymph node (SLN) metastasis in women with apparent uterine-confined endometrial cancer. METHODS: All women underwent SLN identification with complete surgical staging. All SLNs were cut perpendicular to the long axis and two adjacent 5 µm sections were cut at each of two levels 50 µm apart. At each level, one slide was stained with hematoxylin and eosin and the other with immunohistochemistry using the AE1/AE3 anti-cytokeratin antibody, as well as one negative control slide for a total of five slides per block. For OSNA analysis, the 2 mm sections of the lymph nodes were homogenized to form a lysate. The lysate was then centrifuged and inserted into the RD 100i instrument where the isothermal amplification of CK19 mRNA was executed. RESULTS: Of the 396 patients included in the retrospective analysis, 214 were in the SU group, and 182 in the OSNA group. Overall 869 SLNs were identified (490 SU, 379 OSNA). Sixty patients exhibited SLN metastasis (34 SU, 26 OSNA). Macrometastasis, micrometastases, and isolated tumor cells (ITC) were 5.1%, 4.1%, and 0.2%, respectively, in the US group, and 2.4%, 6.3%, and 0.1%, respectively, in the OSNA group (p=0.022). CONCLUSIONS: The OSNA assay detected a higher rate of micrometastasis and a lower rate of macrometastasis and ITC when compared with SU. The clinical and prognostic impact of ITC is debatable and controversial. Further studies are needed to clarify the respective roles of the OSNA and SU methods, and the possible role of ITC in the prognosis of patients with apparent early-stage endometrial cancer.

Role of one-step nucleic acid amplification (OSNA) to detect sentinel lymph node low-volume metastasis in early-stage cervical cancer.

INTRODUCTION: Growing evidence in the literature supports the accuracy of sentinel lymph node (SLN) biopsy in early-stage cervical cancer. One-step nucleic acid amplification (OSNA) is a rapid assay able to detect cytokeratin 19-mRNA in SLNs, and it can be used for intra-operative detection of low-volume metastases. The aim of this study was to evaluate the rate of low-volume metastasis in SLNs detected by OSNA in patients with early-stage cervical cancer. Secondary aims were to define the sensitivity and the negative predictive value of SLN biopsy assessed with OSNA. METHODS: After IRB approval, consecutive patients who underwent surgery for International Federation of Gynecology and Obstetrics stage IA1 with lymph-vascular space involvement to IB1 between November 2017 and July 2019 and had SLN biopsy and pelvic lymphadenectomy were included. SLNs were detected with indocyanine-green cervical injection and sent intra-operatively for OSNA. RESULTS: Eighteen patients underwent SLN assessment with OSNA and systematic pelvic lymphadenectomy in the study period. Four (22.2%) patients had unilateral and 14 (77.8%) had bilateral mapping. OSNA detected micro-metastasis in 6/18 (33.3%) patients. All micro-metastases were detected in patients with bilateral SLN mapping. The sensitivity and negative predictive value of SLN in detecting lymph node metastasis with OSNA calculated per pelvic sidewall were 85.7% and 96.1%, respectively. The false negative rate in mapped sidewalls was 14.3%. DISCUSSION: This is the first series entirely processing SLNs for OSNA in early-stage cervical cancer. OSNA is able to intra-operatively detect low-volume metastases in SLNs. Further studies are necessary to confirm the accuracy of this technique and to assess survival implications of low-volume metastases detected by OSNA.

Nuclear CK19-immunopositive pseudoinclusions as a new additional objective diagnostic feature of papillary thyroid carcinoma.

One of the key parameters in the diagnosis of papillary thyroid carcinoma (PTC) are true nuclear pseudoinclusions (NPs), which constitute invaginations of the cytoplasm into the nucleus. On the other hand, strong cytoplasmic expression of CK19 is a well-known attribute of PTC tumor cells. We analyzed NPs using CK19 immunohistochemistry in histological sections of 52 PTCs and seven noninvasive follicular thyroid neoplasms with papillary-like nuclear features (NIFTPs). Strong CK19+ NPs were present in 77% of PTCs, whereas NPs in hematoxylin and eosin (HE)-stained slides (HE NPs) were identified in only 48% of PTCs. Detection of CK19+ NPs enabled easier and objective recognition of NPs and better discrimination of NPs from pseudo-pseudoinclusions than detection of HE NPs. In the 15 of the 27 (55.5%) PTCs in which we could not discern HE NPs, strong CK19+ NPs could be identified reliably, quickly and easily. Moreover, all NIFTPs were negative for both CK19+ NPs and HE NPs. Detection of CK19+ NPs may refine the assessment of this important diagnostic feature and, hence, the microscopic diagnostic criteria of PTC. Thus, these findings may have implications for the accurate diagnosis of PTC and NIFTP.

Diagnostic and Prognostic Potential of Biomarkers CYFRA 21.1, ERCC1, p53, FGFR3 and TATI in Bladder Cancers.

The high occurrence of bladder cancer and its tendency to recur in combination with a lifelong surveillance make the treatment of superficial bladder cancer one of the most expensive and time-consuming. Moreover, carcinoma in situ often leads to muscle invasion with an unfavorable prognosis. Currently, invasive methods including cystoscopy and cytology remain a gold standard. The aim of this study was to explore urine-based biomarkers to find the one with the best specificity and sensitivity, which would allow optimizing the treatment plan. In this review, we sum up the current knowledge about Cytokeratin fragments (CYFRA 21.1), Excision Repair Cross-Complementation 1 (ERCC1), Tumour Protein p53 (Tp53), Fibroblast Growth Factor Receptor 3 (FGFR3), Tumor-Associated Trypsin Inhibitor (TATI) and their potential applications in clinical practice.

CYFRA 21-1 and Placental Growth Factor as Screening Markers for Endometriosis.

BACKGROUND This study evaluated the performance of serum CYFRA 21-1 and placental growth factor (PIGF) as screening markers for endometriosis. MATERIAL AND METHODS In this prospective study included 81 female patients who underwent laparoscopy to treat benign ovarian tumors. Serum samples were obtained from all study patients before surgery. Serum marker levels, including CYFRA 21-1, PIGF, cancer antigen (CA)125, CA19-9, and human epididymis protein 4 (HE4) were measured using a fluorescence immunoassay technique. RESULTS Forty of the patients were diagnosed with endometriosis (the study group) and 41 women were diagnosed with other benign ovarian tumors (the control group). Mean serum CYFRA 21-1 and PIGF levels were not different between these 2 groups (P=0.179 and P=0.865, respectively). Elevated serum CA125 levels (>35 U/mL) and lower CYFRA 21-1 levels (≤2.29 ng/mL) were more frequently observed in the endometriosis study group than in the control group (P<0.0001, and P=048, respectively). High serum PIGF levels (>14.2 pg/mL) were observed in both groups (P=0.226). Mean serum CA19-9 levels and HE4 levels, as well as the ROMA (risk of ovarian malignancy Algorithm) score were similar between the 2 groups. Sensitivity (95.0%) and negative predictive value (NPV) (80.0%) of CYFRA 21-1 for diagnosing endometriosis were higher than those of CA125 (sensitivity 67.5%, NPV 74.5%) and PIGF (sensitivity 20.0%, NPV 53.6%). However, the specificity (PIGF 90.2%, CA125 92.7%) and positive predictive value (PPV) (PIGF 66.7%, CA125 87.1%) of PIGF and CA125 for diagnosing endometriosis were higher than those of CYFRA 21-1 (specificity 19.5%, PPV 53.5%). CONCLUSIONS CYFRA 21-1 and PIGF may be promising markers to identify patients with and without ovarian endometriosis.

Electrochemiluminescent immunoassay for the lung cancer biomarker CYFRA21-1 using MoOx quantum dots.

Molybdenum oxide quantum dots (MoOx QDs) were synthesized by a one-pot method and used as a versatile probe in an electrochemiluminescent (ECL) immunoassay of the non-small cell lung cancer biomarker cytokeratin 19 fragment 21-1 (CYFRA21-1) as a model analyte. The MoOx QDs exhibited stable and strong cathodic green ECL, with an emission peak at 535 nm, in the presence of K2S2O8 within the potential range of -2.0 to 0 V. On exposure to CYFRA21-1, the ECL decreases because of the immunoreaction between CYFRA21-1 and its antibody which generates a barrier for electron transfer. The determination of CYFRA21-1 with favorable analytical performances was successfully realized under the optimal conditions. ECL decreases linearly in the 1 pg mL-1 to 350 ng mL-1 CYFRA21-1 concentration range, and the detection is as low as 0.3 pg mL-1. Excellent recoveries from CYFRA21-1-spiked human serum indicate that the assay can be operated under physiological conditions. Graphical abstractSchematic representation of the fabrication of molybdenum oxide quantum dots (MoOx QDs) and the electrochemiluminescent (ECL) immunoassay based on the use of the MoOx QDs ECL probe for cytokeratin 19 fragment 21-1 (CYFRA21-1).

Imaging Features of Gadoxetic Acid-enhanced and Diffusion-weighted MR Imaging for Identifying Cytokeratin 19-positive Hepatocellular Carcinoma: A Retrospective Observational Study.

Purpose To determine the preoperative magnetic resonance (MR) imaging findings potentially most useful for predicting cytokeratin 19 (CK19)-positive hepatocellular carcinoma (HCC) and to evaluate the prognosis after curative resection in patients with a single HCC lesion positive for CK19 compared with patients with HCC who are negative for CK19. Materials and Methods The institutional review board approved this study and waived the requirement for informed consent. Two hundred four patients with CK19-negative HCC and 38 with CK19-positive HCC who underwent curative resection after gadoxetic acid-enhanced and diffusion-weighted MR imaging were retrospectively evaluated in a single institution. Two radiologists evaluated preoperative findings at MR imaging. Significant findings for differentiating the two groups were identified at univariate and multivariate analyses. By using receiver operating characteristic analysis, the optimal cut-off values for quantitative variables were determined. Recurrence-free survival rates after surgery were also compared between groups. Results At multivariate analysis, irregular tumor margin (P = .024), arterial rim enhancement (P < .001), lower tumor-to-liver signal intensity (SI) ratio on hepatobiliary phase (HBP) images (≤0.522; P = .01), and lower tumor-to-liver apparent diffusion coefficient (ADC) ratio (≤0.820; P < .001) were independent significant factors to predict CK19-positive HCC. When three of these four criteria were combined, 63.2% (24 of 38; 95% confidence interval: 46.0%, 78.2%) of CK19-positive HCCs were identified with a specificity of 90.7% (185 of 204; 95% confidence interval: 46.0%, 78.2%). When all four criteria were satisfied, specificity was 99.5% (203 of 204; 95% confidence interval: 97.3%, 100%). Recurrence-free survival rates were significantly lower in patients with CK19-positive HCCs compared with those with CK19-negative HCCs after curative resection (63.9% vs 90.0% at 1 year, 63.9% vs 79.9% at 2 years, and 54.8% vs 70.2% at 3 years, P = .001 by log-rank test). Conclusion At gadoxetic acid-enhanced and diffusion-weighted MR imaging, irregular margin, arterial phase rim enhancement, lower tumor-to-liver ADC ratio, and lower tumor-to-liver SI ratio at HBP imaging may be helpful to predict CK19-positive HCC with early recurrence (<2 years) after curative resection. © RSNA, 2017 Online supplemental material is available for this article.