@misc{HiemannHilgerMicheletal., author = {Hiemann, Rico and Hilger, Nadja and Michel, J{\"o}rg and Nitschke, J{\"o}rg and B{\"o}hm, Alexander and Anderer, Ursula and Weigert, Martin and Sack, Ulrich}, title = {Automatic Analysis of Immunofluorescence Patterns of HEp-2 Cells}, series = {Annals of the New York Academy of Sciences}, volume = {1109}, journal = {Annals of the New York Academy of Sciences}, number = {1}, issn = {0077-8923}, doi = {10.1196/annals.1398.042}, pages = {358 -- 371}, abstract = {The standard screening test for the recognition of autoimmune diseases is the proof of autoantibodies in serum of patients by indirect immunofluorescence (IIF) based on HEp-2 cells. Manual evaluation of this test is very subjective, slow, and there are no objective parameters as guidelines available. Interlaboratory tests showed occasionally large deviations in the test evaluation resulting in a high variance of results. The aim of this project is fast, objective, safe, and economical automatic analysis of HEp-2 IIF patterns. Images of IIF patterns were completely and automatically captured using an inverse motorized fluorescence microscope. Thereby, device-specific parameters were controlled automatically, too. For fast analysis of IIF patterns new algorithms of image processing were developed. Artifacts were recognized and excluded from analysis by the developed software. Analysis of more than 80,000 images clearly demonstrated full automatization and fast processing of IIF patterns. Additionally serum-specific fluorescence could be easily distinguished from background. Even very weak but positive patterns can be recognized and used for diagnosis. A detailed separation into different basic patterns is possible. Objective, fast, and disease-related economical analysis of HEp-2 immunofluorescence patterns is feasible. The implemented software algorithms allowed a mathematical way of describing IIF patterns and can therefore be a useful tool for the needed standardization process.}, language = {en} } @misc{HiemannRoggenbuckSacketal., author = {Hiemann, Rico and Roggenbuck, Dirk and Sack, Ulrich and Anderer, Ursula and Conrad, Karsten}, title = {Die HEp-2-Zelle als Target f{\"u}r multiparametrische Autoantik{\"o}rperanalytik - Automatisierung und Standardisierung/The HEp-2 cell as target for multiparametric autoantibody analyses: automation and standardisation}, series = {Journal of Laboratory Medicine}, volume = {35}, journal = {Journal of Laboratory Medicine}, number = {6}, issn = {1439-0477}, doi = {10.1515/JLM.2011.053}, pages = {351 -- 361}, abstract = {Trotz Einf{\"u}hrung von Multiplex-Assays ist die Bestimmung von Autoantik{\"o}rpern (AAK) mittels indirekter Immunfluoreszenz (IIF) nach wie vor ein wesentlicher Bestandteil der Autoimmundiagnostik. Das Screening auf nichtorganspezifische AAK wird in der Routinediagnostik von autoimmunen rheumatischen Erkrankungen mittels indirekter Immunfluoreszenz an HEp-2-Zellen durchgef{\"u}hrt. Leider differieren die mit diesem Test erhaltenen Befunde z. T. recht erheblich auf Grund von objektiven (z. B. HEp-2-Zellfixierung) und subjektiven (z. B. Erfahrungswerte) Faktoren. Die Intra- und Interlaborvarianzen sind daher relativ hoch. Erste Ergebnisse von Evaluierungsstudien eines neu entwickelten Systems zur automatischen Bildanalyse (AKLIDES®) zeigen einen Weg in Richtung Standardisierung (bzw. Harmonisierung) des HEp-2-Zell-Assays auf, in dem zun{\"a}chst subjektive Fehlinterpretationen vermieden werden k{\"o}nnen. Eine weitere Harmonisierung erfordert die Weiterentwicklung der Software-Algorithmen zur Mustererkennung sowie neue Kalibrationssysteme.}, language = {en} } @misc{LiedtkeRoseHiemannetal., author = {Liedtke, Victoria and Rose, Laura and Hiemann, Rico and Nasser, Abdullah and R{\"o}diger, Stefan and Bonaventura, Alena and Winkler, Laura and Sowa, Mandy and St{\"o}ckle, Michael and Schierack, Peter and Junker, Kerstin and Roggenbuck, Dirk}, title = {Over-Expression of LEDGF/p75 in HEp-2 Cells Enhances Autoimmune IgG Response in Patients with Benign Prostatic Hyperplasia—A Novel Diagnostic Approach with Therapeutic Consequence?}, series = {International Journal of Molecular Sciences}, volume = {24}, journal = {International Journal of Molecular Sciences}, number = {7}, issn = {1422-0067}, doi = {10.3390/ijms24076166}, abstract = {Lens epithelium-derived growth factor splice variant of 75 kDa (LEDGF/p75) is an autoantigen over-expressed in solid tumors and acts as a stress-related transcriptional co-activator. Participation of autoimmune responses in the pathophysiology of benign prostatic hyperplasia (PBH) and a corresponding immunosuppressive therapy by TNFalpha antagonists has been recently suggested. Thus, autoAb testing could aid in the diagnosis of BPH patients profiting from such therapy. We generated CRISPR/Cas9 modified HEp-2 LEDGF knock-out (KO) and HEp-2 LEDGF/p75 over-expressing (OE) cells and examined IgG autoantibody reactivity to LEDGF/p75 in patients with prostate cancer (PCa, n = 89), bladder cancer (BCa, n = 116), benign prostatic hyperplasia (BPH, n = 103), and blood donors (BD, n = 60) by indirect immunofluorescence assay (IFA). Surprisingly, we could not detect elevated binding of autoAbs against LEDGF/p75 in cancer patients, but autoAb reactivity to LEDGF/p75 OE cells in about 50\% of patients with BPH was unexpectedly significantly increased. Furthermore, a line immunoassay enabling the detection of 18 different autoAbs revealed a significantly increased occurrence of anti-dsDNA autoAbs in 34\% of BPH patients in contrast to tumor patients and BD. This finding was confirmed by anti-mitochondrial (mDNA) autoAb detection with the Crithidia luciliae immunofluorescence test, which also showed a significantly higher prevalence (34\%) of anti-mDNA autoAbs in BPH. In summary, our study provided further evidence for the occurrence of autoimmune responses in BPH. Furthermore, LEDGF/p75 over-expression renders HEp-2 cells more autoantigenic and an ideal target for autoAb analysis in BPH with a potential therapy consequence.}, language = {en} }