The utility of a real-time RT-PCR assay for the detection of metastases greater than 0.2 mm in sentinel lymph nodes of breast cancer patients confirmed by detailed histological analysis

2009 ◽  
Vol 27 (15_suppl) ◽  
pp. 628-628
Author(s):  
M. Kurosumi ◽  
Y. Kobayashi ◽  
H. Takei
Keyword(s):  
Lymph Nodes ◽  
Real Time ◽  
Histological Analysis ◽  
Sentinel Lymph Nodes ◽  
Breast Cancer Patients ◽  
Frozen Sections ◽  
Rt Pcr ◽  
Pcr Assay ◽  
The Real ◽  
Cutoff Values

628 Background: Analysis using real time RT-PCR for the detection of metastases in lymph nodes (LN) increases sampling but is associated with a risk of too much sensitivity as compared to histological analysis. Determining the appropriate cutoff value is important in introducing this system to routine clinical practice. Our study confirms the reliability of the cutoff values of a real-time RT-PCR assay (GeneSearch, Veridex LLC) to detect metastases larger than 0.2 mm by detailed 0.2 mm frozen section histological diagnosis. Methods: 129 sentinel and non-sentinel lymph nodes were obtained by sentinel LN biopsy from 79 breast cancer patients. All LNs were cut in half. One half of each LN was used for routine intra-operative diagnosis. The other LN half had sections cut every 0.2 mm for histological analysis. All frozen sections were H&E stained and examined by on-site pathologists. These sections were then re-stained immunohistochemically using a pancytokeratin antibody (AE1/AE3) for detecting submicrometastses. All remaining tissue between the histology sections was assayed by the real-time RT-PCR assay using the genetic markers mammaglobin (MG) and cytokeratin-19 (CK-19) (note: tissue processing method was investigational and off-label). Cutoff values were pre-set in a large US study (n = 304). Results: Compared to the histological diagnosis using 0.2 mm interval frozen sections, the real-time RT-PCR results were as follows; sensitivity of 100.0% (34/34), specificity of 93.7% (89/95), and overall accuracy of 95.3% (123/129). Submicrometastases were recognized by IHC in 3 of 95 samples (2 solitary and 1 multiple sites) diagnosed as negative by H&E-stain sections, and 3 samples were negative for RT-PCR. Conclusions: Results of 0.2 mm interval histological analysis suggest a near perfect sensitivity of the real time RT-PCR assay, allowing reliable detection of LN metastases larger than 0.2 mm. In addition, at least three samples with submicrometastases detected by immunohistochemistry for pancytokeratin were above the preset cutoff values of this real time RT-PCR assay. [Table: see text]

The utility of an investigational real-time RT-PCR assay system for the detection of micrometastases in sentinel lymph nodes of breast cancer confirmed by 0.2 mm histological interval analysis

2007 ◽  
Vol 25 (18_suppl) ◽  
pp. 10560-10560
Author(s):  
M. Kurozumi ◽  
Y. Kobayashi ◽  
H. Takei ◽  
K. Kitsugi ◽  
M. Ueno ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Lymph Node ◽  
Real Time ◽  
Interval Analysis ◽  
Histological Analysis ◽  
Frozen Section ◽  
Breast Cancer Patients ◽  
Rt Pcr ◽  
Pcr Assay ◽  
The Real

10560 Background: Comparison between real-time RT-PCR and histological analysis for the detection of lymph node metastasis in breast cancer patients is difficult because of the necessary use of different tissue for the two tests. In order to minimize this challenge this study used histology sections taken every 0.2 mm and all tissue between the sections was analyzed by real-time RT-PCR. Methods: 129 Lymph nodes (LNs) were obtained by Sentinel LN biopsy and/or axillary dissection from 80 breast cancer patients. All LNs were cut in half. One half of each LN was used for the routine intra-operative diagnosis. The other half was used for the 0.2mm histological interval analysis. All 10μm sections cut for histology at 0.2 mm intervals were analyzed using frozen section H&E under a light microscope. All remaining tissue between the sections used for histological analysis was assayed by real-time RT-PCR using the genetic markers mammaglobin and cytokeratin-19 (investigational GeneSearch Breast Lymph Node Assay, Veridex, LLC, Warren, NJ, USA). Cutoffs were pre- set for the real-time RT-PCR to detect only metastasis greater than 0.2 mm. Results: Compared to the histological diagnosis using 0.2 mm interval frozen sections, the real-time RT-PCR results were as follows; sensitivity of 100.0% (34/34), specificity of 93.7% (89/95) and overall accuracy of 95.3% (123/129). Conclusions: These data suggest by utilizing the 0.2 mm histological interval analysis, sampling discrepancies are minimized and a high level of sensitivity and overall accuracy is seen for the real-time RT-PCR assay. Assay specificity calculations may be less than 100% due to interpretation challenges in frozen section histology, especially in the cases of smaller metastases and/or lobular cancer, causing a small percentage of clinically relevant metastasis to be missed. [Table: see text]

Assessing the status of axillary sentinel lymph nodes of breast carcinoma patients by a real-time quantitative RT-PCR assay for mammaglobin 1 mRNA

2006 ◽  
Vol 98 (2) ◽  
pp. 185-190 ◽  
Author(s):  
Patrizia Dell’Orto ◽  
Maria Olivia Biasi ◽  
Barbara Del Curto ◽  
Stefano Zurrida ◽  
Viviana Galimberti ◽  
...  
Keyword(s):  
Breast Carcinoma ◽  
Lymph Nodes ◽  
Real Time ◽  
Sentinel Lymph Nodes ◽  
Rt Pcr ◽  
Pcr Assay ◽  
The Status

scholarly journals Description and Validation of a Novel Real-Time RT-PCR Enterovirus Assay

2008 ◽  
Vol 54 (2) ◽  
pp. 406-413 ◽  
Author(s):  
Weston C Hymas ◽  
Wade K Aldous ◽  
Edward W Taggart ◽  
Jeffery B Stevenson ◽  
David R Hillyard
Keyword(s):  
Sequence Analysis ◽  
Single Nucleotide Polymorphisms ◽  
Real Time ◽  
Clinical Samples ◽  
Nucleotide Polymorphisms ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Single Nucleotide ◽  
The Real ◽  
Nontranslated Region

Abstract Background: Enteroviruses are a leading cause of aseptic meningitis in adult and pediatric populations. We describe the development of a real-time RT-PCR assay that amplifies a small target in the 5′ nontranslated region upstream of the classical Rotbart enterovirus amplicon. The assay includes an RNA internal control and incorporates modified nucleotide chemistry. Methods: We evaluated the performance characteristics of this design and performed blinded parallel testing on clinical samples, comparing the results with a commercially available RT-PCR assay (Pan-Enterovirus OligoDetect kit) that uses an enzyme immunoassay–like plate end detection. Results: We tested 778 samples and found 14 discrepant samples between the 2 assays. Of these, the real-time assay detected 6 samples that were negative by the OligoDetect kit, 5 of which were confirmed as positive by sequence analysis using an alternative primer set. Eight discrepant samples were positive by the OligoDetect kit and real-time negative, with 6 confirmed by sequencing. Overall, detection rates of 97% and 96% were obtained for the OligoDetect kit and real-time assays, respectively. Sequence analysis revealed the presence of a number of single nucleotide polymorphisms in the targeted region. The comparative sensitivities of the 2 assays were equivalent, with the limit of detection for the real-time assay determined to be approximately 430 copies per milliliter in cerebrospinal fluid. Conclusions: This novel real-time enterovirus assay is a sensitive and suitable assay for routine clinical testing. The presence of single nucleotide polymorphisms can affect real-time PCR assays.

scholarly journals Rapid SARS-CoV-2 antigen detection assay in comparison with real-time RT-PCR assay for laboratory diagnosis of COVID-19 in Thailand

2020 ◽  
Vol 17 (1) ◽  
Author(s):  
Chutikarn Chaimayo ◽  
Bualan Kaewnaphan ◽  
Nattaya Tanlieng ◽  
Niracha Athipanyasilp ◽  
Rujipas Sirijatuphat ◽  
...  
Keyword(s):  
Real Time ◽  
Sensitivity And Specificity ◽  
False Negative ◽  
Laboratory Diagnosis ◽  
Antigen Detection ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Screening Assay ◽  
The Real ◽  
Pcr Test

Abstract Background The Coronavirus disease 2019 (COVID-19) pandemic continues to spread across the world. Hence, there is an urgent need for rapid, simple, and accurate tests to diagnose severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Performance characteristics of the rapid SARS-CoV-2 antigen detection test should be evaluated and compared with the gold standard real-time reverse transcription-polymerase chain reaction (RT-PCR) test for diagnosis of COVID-19 cases. Methods The rapid SARS-CoV-2 antigen detection test, Standard™ Q COVID-19 Ag kit (SD Biosensor®, Republic of Korea), was compared with the real-time RT-PCR test, Allplex™ 2019-nCoV Assay (Seegene®, Korea) for detection of SARS-CoV-2 in respiratory specimens. Four hundred fifty-four respiratory samples (mainly nasopharyngeal and throat swabs) were obtained from COVID-19 suspected cases and contact individuals, including pre-operative patients at Siriraj Hospital, Bangkok, Thailand during March–May 2020. Results Of 454 respiratory samples, 60 (13.2%) were positive, and 394 (86.8%) were negative for SARS-CoV-2 RNA by real-time RT-PCR assay. The duration from onset to laboratory test in COVID-19 suspected cases and contact individuals ranged from 0 to 14 days with a median of 3 days. The rapid SARS-CoV-2 antigen detection test’s sensitivity and specificity were 98.33% (95% CI, 91.06–99.96%) and 98.73% (95% CI, 97.06–99.59%), respectively. One false negative test result was from a sample with a high real-time RT-PCR cycle threshold (Ct), while five false positive test results were from specimens of pre-operative patients. Conclusions The rapid assay for SARS-CoV-2 antigen detection showed comparable sensitivity and specificity with the real-time RT-PCR assay. Thus, there is a potential use of this rapid and simple SARS-CoV-2 antigen detection test as a screening assay.

Frozen Sections of Sentinel Lymph Nodes From Breast Cancer Patients Who Undergo Neoadjuvant Therapy Are Accurately Diagnosed Using Telepathology

2019 ◽  
Vol 152 (Supplement_1) ◽  
pp. S122-S123
Author(s):  
Valarie McMurtry ◽  
Rachel Factor
Keyword(s):  
Breast Cancer ◽  
Lymph Nodes ◽  
Neoadjuvant Therapy ◽  
Cancer Patients ◽  
Treatment Effect ◽  
Frozen Section ◽  
Lobular Carcinoma ◽  
Sentinel Lymph Nodes ◽  
Breast Cancer Patients ◽  
Frozen Sections

Abstract Objectives Telepathology enables histologic diagnosis to be made from a scanned slide visualized on a computer. Frozen sections (FSs) can be performed at remote locations and read by a pathologist at a central site. At our institution, qualifying breast cancer patients are enrolled in clinical trials that require FS on sentinel lymph nodes (SLNs) after neoadjuvant therapy (NAT), including chemotherapy or endocrine therapy. In this setting, histology is complicated by treatment effect and biopsy site changes. Others have reported good accuracy of FSs on SLNs after NAT. We investigated whether pathologists are accurate in diagnosing SLN FSs for such cases while using telepathology. To our knowledge, this has not been reported previously. Methods SLNs were entirely submitted and serially sectioned (2-mm thickness). At least two levels were cut. All FSs were submitted for formalin-fixed, paraffin-embedded permanent sections. A pathology assistant at the remote location prepared the FSs and scanned slides using the VisionTek M6 digital microscope ecosystem (East Dundee, IL). Cases were interpreted by board-certified pathologists who completed training on the VisionTek system. For this study, diagnoses from FSs and permanents were compared. Results Forty-five SLNs from 19 breast neoadjuvant cases were read by VisionTek from March 2017 to January 2019. Forty-three cases (96%) called negative by FSs were confirmed negative (on permanents). One hundred percent called positive by FSs were positive. Four SLN called atypical on FSs were positive. Three of these were neoadjuvant endocrine cases for invasive lobular carcinoma. Two cases called atypical were negative. One of these, called atypical/suspicious, resulted in axillary dissection. This case was reviewed by three pathologists at the time of surgery. It had abundant treatment effect, mimicking carcinoma. Conclusion While pitfalls exist, overall, the diagnostic accuracy of frozen section analysis by telepathology of SLNs from breast cases after neoadjuvant therapy is high.

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