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Stakenborg T.,IMEC | Liu C.,IMEC | Henry O.,Rovira i Virgili University | O'Sullivan C.K.,Rovira i Virgili University | And 7 more authors.
2010 Annual International Conference of the IEEE Engineering in Medicine and Biology Society, EMBC'10 | Year: 2010

A smart miniaturized system is being proposed for the isolation and characterization of circulating tumor cells (CTCs) directly from blood. Different microfluidic modules have been designed for cell enrichment and -counting, multiplex mRNA amplification as well as DNA detection. With the different modules at hand, future effort will focus on the integration of the modules in a fully automated, single platform. © 2010 IEEE.


Stakenborg T.,IMEC | Liu C.,IMEC | Henry O.,Rovira i Virgili University | Borgen E.,University of Oslo | And 7 more authors.
Expert Review of Molecular Diagnostics | Year: 2010

Cancer remains a prominent health concern in modern societies. Continuous innovations and introduction of new technologies are essential to level or reduce current healthcare spending. A diagnostic platform to detect circulating tumor cells (CTCs) in peripheral blood may be most promising in this respect. CTCs have been proposed as a minimally invasive, prognostic and predictive marker to reflect the biological characteristics of tumors and are implemented in an increasing number of clinical studies. Still, their detection remains a challenge as they may occur at concentrations below one single cell per ml of blood. To facilitate their detection, here we describe microfluidic modules to isolate and genotype CTCs directly from clinical blood samples. In a first cell isolation and detection module, the CTCs are immunomagnetically enriched, separated and counted. In a second module and after cell lysis, the mRNA is reversely transcripted to cDNA, followed by a multiplex ligation probe amplification of 20 specific genetic markers and two control fragments. Following the multiplex ligation probe amplification reaction, the amplified fragments are electrochemically detected in a third and final module. Besides the design of the modules, their functionality is described using control samples. Further testing using clinical samples and integration of all modules in a single, fully automated smart miniaturized system will enable minimal invasive testing for frequent detection and characterization of CTCs. © 2010 Expert Reviews Ltd.


Kuhlmann J.D.,University of Duisburg - Essen | Kuhlmann J.D.,TU Dresden | Kuhlmann J.D.,German Cancer Research Center | Wimberger P.,University of Duisburg - Essen | And 16 more authors.
Clinical Chemistry | Year: 2014

BACKGROUND: Platinum resistance constitutes one of the most recognized clinical challenges for ovarian cancer. Notably, the detection of the primary tumor-based excision repair cross-complementation group 1 (ERCC1) protein by immunohistochemistry was recently shown to be inaccurate for the prediction of platinum resistance. On the basis of the previous finding that circulating tumor cells (CTC) in the blood of ovarian cancer patients are prognostically significant, and given our hypothesis that the negative prognostic impact of CTC may arise from a cellular phenotype associated with platinum resistance, we asked whether expression of the excision repair cross-complementation group 1 (ERCC1) gene in the form of the ERCC1 transcript in CTC may be a suitable blood-based biomarker for platinum resistance. METHODS: The presence of CTC was analyzed by immunomagnetic CTC enrichment (n = 143 patients) targeting the epithelial epitopes epithelial cell adhesion molecule (EPCAM) (also known as GA733-2) and mucin 1, cell surface associated (MUC1), followed by multiplex reverse-transcription PCR to detect the transcripts EPCAM, MUC1, and mucin 16, cell surface associated (MUC16) (also known as CA125), including ERCC1 transcripts in a separate approach. ERCC1 expression in primary tumors was comparatively assessed by immunohistochemistry, using the antibody 8F1. RESULTS: At primary diagnosis, the presence of CTC was observed in 14% of patients and constituted an independent predictor of overall survival (OS) (P = 0.041). ERCC1-positive CTC (ERCC1+CTC) were observed in 8% of patients and constituted an independent predictor, not only for OS but also for progression-free survival (PFS) (P = 0.026 and P = 0.009, respectively). More interestingly, we discovered the presence of ERCC1+CTC at primary diagnosis to be likewise an independent predictor of platinum resistance (P = 0.010), whereas ERCC1 expression in corresponding primary tumor tissue predicted neither platinum resistance nor prognosis. CONCLUSIONS: The presence of ERCC1+CTC can serve as a blood-based diagnostic biomarker for predicting platinum resistance at primary diagnosis of ovarian cancer. © 2014 American Association for Clinical Chemistry.


PubMed | German Cancer Research Center, University of Duisburg - Essen, Adnagen Inc. and Acomed Statistics
Type: Journal Article | Journal: Clinical chemistry | Year: 2014

Platinum resistance constitutes one of the most recognized clinical challenges for ovarian cancer. Notably, the detection of the primary tumor-based excision repair cross-complementation group 1 (ERCC1) protein by immunohistochemistry was recently shown to be inaccurate for the prediction of platinum resistance. On the basis of the previous finding that circulating tumor cells (CTC) in the blood of ovarian cancer patients are prognostically significant, and given our hypothesis that the negative prognostic impact of CTC may arise from a cellular phenotype associated with platinum resistance, we asked whether expression of the excision repair cross-complementation group 1 (ERCC1) gene in the form of the ERCC1 transcript in CTC may be a suitable blood-based biomarker for platinum resistance.The presence of CTC was analyzed by immunomagnetic CTC enrichment (n = 143 patients) targeting the epithelial epitopes epithelial cell adhesion molecule (EPCAM) (also known as GA733-2) and mucin 1, cell surface associated (MUC1), followed by multiplex reverse-transcription PCR to detect the transcripts EPCAM, MUC1, and mucin 16, cell surface associated (MUC16) (also known as CA125), including ERCC1 transcripts in a separate approach. ERCC1 expression in primary tumors was comparatively assessed by immunohistochemistry, using the antibody 8F1.At primary diagnosis, the presence of CTC was observed in 14% of patients and constituted an independent predictor of overall survival (OS) (P = 0.041). ERCC1-positive CTC (ERCC1(+)CTC) were observed in 8% of patients and constituted an independent predictor, not only for OS but also for progression-free survival (PFS) (P = 0.026 and P = 0.009, respectively). More interestingly, we discovered the presence of ERCC1(+)CTC at primary diagnosis to be likewise an independent predictor of platinum resistance (P = 0.010), whereas ERCC1 expression in corresponding primary tumor tissue predicted neither platinum resistance nor prognosis.The presence of ERCC1(+)CTC can serve as a blood-based diagnostic biomarker for predicting platinum resistance at primary diagnosis of ovarian cancer.


Acero Sanchez J.L.,Rovira i Virgili University | Henry O.Y.F.,Rovira i Virgili University | Mairal T.,Rovira i Virgili University | Laddach N.,MRC Holland | And 5 more authors.
Analytical and Bioanalytical Chemistry | Year: 2010

An enzyme-linked oligonucleotide assay (ELONA) for quantification of mRNA expression of five genes involved in breast cancer, extracted from isolated rare tumour cells and amplified by multiplex ligation-dependent probe amplification (MLPA) is presented. In MLPA, a multiplex oligonucleotide ligation assay is combined with a PCR reaction in which all ligation products are amplified by use of a single primer pair. Biotinylated probes complementary to each of the target sequences were immobilised on the surface of a streptavidin-coated microtitre plate and exposed to single-stranded MLPA products. A universal reporting probe sequence modified with horseradish peroxidase (URP-HRP) and complementary to a universal primer used during the MLPA step was further added to the surface-bound duplex as a reporter probe. Simultaneous addition of anchoring probe and target, followed by addition of reporter probe, rather than sequential addition, was achieved with no significant effect on sensitivity and limits of detection, but considerably reduced the required assay time. Detection limits as low as 20 pmol L-1, with an overall assay time of 95 min could be achieved with negligible cross-reactivity between probes and non-specific targets present in the MLPA-PCR product. The same MLPA-PCR product was analysed using capillary electrophoresis, the technique typically used for analysis of MLPA products, and good correlation was observed. The assay presented is easy to carry out, relatively inexpensive, rapid, does not require sophisticated instrumentation, and enables quantitative analysis, making it very promising for the analysis of MLPA products. © 2010 Springer-Verlag.


The present invention concerns a monoclonal antibody and corresponding hybridoma cells and antigens suitable for isolating fetal cells from maternal blood. The inventive monoclonal antibody reacts with a surface antigen present on fetal red blood cells including their nucleated precursor cells, but not with surface antigens on adult erythroid cell.


Adnagen Inc. | Entity website


Adnagen Inc. | Entity website


Adnagen Inc. | Entity website

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