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Wagner, Niklas ; Rapp, Anna Elise ; Braun, Sebastian Benedict ; Ehnert, Markus ; Imhof, Thomas ; Koch, Manuel ; Jenei-Lanzl, Zsuzsa ; Zaucke, Frank ; Meurer, Andrea
Matrix metalloproteinases (MMPs) play crucial roles in tissue homeostasis and pathologies by remodeling the extracellular matrix. Previous studies have demonstrated the biological activities of MMP-derived cleavage products. Furthermore, specific fragments can serve as biomarkers. Therefore, an in vitro cleavage assay to identify substrates and characterize cleavage patterns could provide important insight in disease-relevant mechanisms and the identification of novel biomarkers. In the pathogenesis of osteoarthritis (OA), MMP-2, -8, -9 and -13 are of vital importance. However, it is unclear which protease can cleave which matrix component. To address this question, we established an in vitro cleavage assay using recombinantly expressed MMPs and the two cartilage matrix components, COMP and thrombospondin-4. We found a time- and concentration-dependent degradation and an MMP-specific cleavage pattern for both proteins. Cleavage products can now be enriched and purified to investigate their biological activity. To verify the in vivo relevance, we compared the in vitro cleavage patterns with serum and synovial fluid from OA patients and could indeed detect fragments of similar size in the human samples. The cleavage assay can be adapted to other MMPs and substrates, making it a valuable tool for many research fields.
Ong, Yu Ting ; Andrade, Jorge ; Armbruster, Max ; Shi, Chenyue ; Castro, Marco ; Costa, Ana S. H. ; Sugino, Toshiya ; Eelen, Guy ; Zimmermann, Barbara ; Wilhelm, Kerstin ; Lim, Joseph ; Watanabe, Shuichi ; Günther, Stefan ; Schneider, Andre ; Zanconato, Francesca ; Kaulich, Manuel ; Pan, Duojia ; Braun, Thomas ; Gerhardt, Holger ; Efeyan, Alejo ; Carmeliet, Peter ; Piccolo, Stefano ; Grosso, Ana Rita ; Potente, Michael
Angiogenesis, the process by which endothelial cells (ECs) form new blood vessels from existing ones, is intimately linked to the tissue’s metabolic milieu and often occurs at nutrient-deficient sites. However, ECs rely on sufficient metabolic resources to support growth and proliferation. How endothelial nutrient acquisition and usage are regulated is unknown. Here we show that these processes are instructed by Yes-associated protein 1 (YAP)/WW domain-containing transcription regulator 1 (WWTR1/TAZ)-transcriptional enhanced associate domain (TEAD): a transcriptional module whose function is highly responsive to changes in the tissue environment. ECs lacking YAP/TAZ or their transcriptional partners, TEAD1, 2 and 4 fail to divide, resulting in stunted vascular growth in mice. Conversely, activation of TAZ, the more abundant paralogue in ECs, boosts proliferation, leading to vascular hyperplasia. We find that YAP/TAZ promote angiogenesis by fuelling nutrient-dependent mTORC1 signalling. By orchestrating the transcription of a repertoire of cell-surface transporters, including the large neutral amino acid transporter SLC7A5, YAP/TAZ-TEAD stimulate the import of amino acids and other essential nutrients, thereby enabling mTORC1 activation. Dissociating mTORC1 from these nutrient inputs—elicited by the loss of Rag GTPases—inhibits mTORC1 activity and prevents YAP/TAZ-dependent vascular growth. Together, these findings define a pivotal role for YAP/TAZ-TEAD in controlling endothelial mTORC1 and illustrate the essentiality of coordinated nutrient fluxes in the vasculature.
Lang, Franziska ; Aravamudhan, Sriram ; Nolte, Hendrik ; Türk, Clara ; Hölper, Soraya ; Müller, Stefan ; Günther, Stefan ; Blaauw, Bert ; Braun, Thomas ; Krüger, Marcus
Loss of neuronal stimulation enhances protein breakdown and reduces protein synthesis, causing rapid loss of muscle mass. To elucidate the pathophysiological adaptations that occur in atrophying muscles, we used stable isotope labelling and mass spectrometry to quantify protein expression changes accurately during denervation-induced atrophy after sciatic nerve section in the mouse gastrocnemius muscle. Additionally, mice were fed a stable isotope labelling of amino acids in cell culture (SILAC) diet containing 13C6-lysine for 4, 7 or 11 days to calculate relative levels of protein synthesis in denervated and control muscles. Ubiquitin remnant peptides (K-ε-GG) were profiled by immunoaffinity enrichment to identify potential substrates of the ubiquitin-proteasomal pathway. Of the 4279 skeletal muscle proteins quantified, 850 were differentially expressed significantly within 2 weeks after denervation compared with control muscles. Moreover, pulse labelling identified Lys6 incorporation in 4786 proteins, of which 43 had differential Lys6 incorporation between control and denervated muscle. Enrichment of diglycine remnants identified 2100 endogenous ubiquitination sites and revealed a metabolic and myofibrillar protein diglycine signature, including myosin heavy chains, myomesins and titin, during denervation. Comparative analysis of these proteomic data sets with known atrogenes using a random forest approach identified 92 proteins subject to atrogene-like regulation that have not previously been associated directly with denervation-induced atrophy. Comparison of protein synthesis and proteomic data indicated that upregulation of specific proteins in response to denervation is mainly achieved by protein stabilization. This study provides the first integrated analysis of protein expression, synthesis and ubiquitin signatures during muscular atrophy in a living animal.
Van Lunzen, Jan ; Fätkenheuer, Gerd ; Lutz, Thomas ; Klauke, Stefan ; Mauß, Stefan ; Knechten, Heribert ; Braun, Patrick ; Gallo, Lothar ; Ranneberg, Britta
Poster presentation: Purpose of the study First-line HAART with tenofovir DF (TDF) and FTC in pivotal trials has been associated with high efficacy and good tolerability. However, real-life clinical practice often differs from clinical trials due to co-morbidities, co-infections, and less intensive clinical monitoring. To evaluate efficacy and safety of first-line HAART in a day-to-day setting, this Gilead-sponsored non-interventional cohort was established. Methods Between July 2005 and August 2006, 533 HIV-1 infected antiretroviral-naïve patients from 50 German centres enrolled in this non-interventional cohort. All patients were followed every 3 months for 3 years to monitor efficacy (viral load [VL], CD4), tolerability, renal safety, regimen changes and resistance profile. All patients received TDF+FTC as a single tablet (Truvada, TVD) in combination with either an NNRTI or PI/r as their first antiretroviral regimen. Summary of results As of June 2008, 2 years of therapy have been documented for 330/533 (62%) patients. At treatment initiation, 81% were male; median age was 39 years; clinical AIDS diagnosis was documented in 22%; 47% started therapy with CD4 <200 cells/mm3. TVD was combined with an NNRTI (43%) or a PI/r (57%). After 24 months, in an As-Treated (AT) analysis, 85% patients achieved a VL <50 copies/ml (VL <500 copies/ml: 97%), median CD4 count increased from 217 at baseline to 450 cells/mm3 (IQR: 325–608). Truvada showed a good safety profile; 76 adverse events (AEs) of any grade were reported in 66/533 patients (12%); six of these were judged serious. Fourteen (2.6%) patients discontinued TVD due to AEs. Renal abnormalities of any grade were reported in 10 patients (1.9%). Virological failure was documented in nine patients, of which eight were genotyped; M184V/I was detected in three, K65R in two patients. Conclusion During 2 years of follow-up, the overall safety of TVD was good; renal AEs of any grade were reported in 1.9% of patients. K65R was detected in two patients. First-line HAART with TVD plus an NNRTI or PI/r in clinical practice showed comparable efficacy to that observed in controlled clinical trials.
Eißmann, Moritz ; Gutschner, Tony ; Hämmerle, Monika ; Günther, Stefan ; Caudron-Herge, Maïwen ; Groß, Matthias ; Schirmacher, Peter ; Rippe, Karsten ; Braun, Thomas ; Diederichs, Sven ; Zörnig, Martin
The metastasis-associated lung adenocarcinoma transcript 1, MALAT1, is a long non-coding RNA (lncRNA) that has been discovered as a marker for lung cancer metastasis. It is highly abundant, its expression is strongly regulated in many tumor entities including lung adenocarcinoma and hepatocellular carcinoma as well as physiological processes, and it is associated with many RNA binding proteins and highly conserved throughout evolution. The nuclear transcript MALAT-1 has been functionally associated with gene regulation and alternative splicing and its regulation has been shown to impact proliferation, apoptosis, migration and invasion.
Here, we have developed a human and a mouse knockout system to study the loss-of-function phenotypes of this important ncRNA. In human tumor cells, MALAT1 expression was abrogated using Zinc Finger Nucleases. Unexpectedly, the quantitative loss of MALAT1 did neither affect proliferation nor cell cycle progression nor nuclear architecture in human lung or liver cancer cells. Moreover, genetic loss of Malat1 in a knockout mouse model did not give rise to any obvious phenotype or histological abnormalities in Malat1-null compared with wild-type animals. Thus, loss of the abundant nuclear long ncRNA MALAT1 is compatible with cell viability and normal development.
Rieke, Johanna Magdalena ; Zhang, Rong ; Braun, Doreen ; Yilmaz, Öznur ; Japp, Anna S. ; Lopes, Filipa M. ; Pleschka, Michael ; Hilger, Alina Christine ; Schneider, Sophia ; Newman, William G. ; Schneider, Sophia ; Newman, William G. ; Beaman, Glenda M. ; Nordenskjöld, Agneta ; Ebert, Anne-Karoline ; Promm, Martin ; Rösch, Wolfgang H. ; Stein, Raimund ; Hirsch, Karin ; Schäfer, Frank-Mattias ; Schmiedeke, Eberhard ; Boemers, Thomas M. ; Lacher, Martin ; Kluth, Dietrich ; Gosemann, Jan-Hendrik ; Anderberg, Magnus ; Barker, Gillian ; Holmdahl, Gundela ; Läckgren, Göran ; Keene, David ; Cervellione, Raimondo M. ; Giorgio, Elisa ; Di Grazia, Massimo ; Feitz, Wouter F. J. ; Marcelis, Carlo L. M. ; Rooij, Iris Antonia Leonarda Martina van ; Bökenkamp, Arend ; Beckers, Goedele M. A. ; Keegan, Catherine E. ; Sharma, Amit ; Dakal, Tikam Chand ; Wittler, Lars ; Grote, Phillip ; Zwink, Nadine ; Jenetzky, Ekkehart ; Brusco, Alfredo ; Thiele, Holger ; Ludwig, Michael ; Schweizer, Ulrich ; Woolf, Adrian S. ; Odermatt, Benjamin ; Reutter, Heiko
Previous studies in developing Xenopus and zebrafish reported that the phosphate transporter slc20a1a is expressed in pronephric kidneys. The recent identification of SLC20A1 as a monoallelic candidate gene for cloacal exstrophy further suggests its involvement in the urinary tract and urorectal development. However, little is known of the functional role of SLC20A1 in urinary tract development. Here, we investigated this using morpholino oligonucleotide knockdown of the zebrafish ortholog slc20a1a. This caused kidney cysts and malformations of the cloaca. Moreover, in morphants we demonstrated dysfunctional voiding and hindgut opening defects mimicking imperforate anus in human cloacal exstrophy. Furthermore, we performed immunohistochemistry of an unaffected 6-week-old human embryo and detected SLC20A1 in the urinary tract and the abdominal midline, structures implicated in the pathogenesis of cloacal exstrophy. Additionally, we resequenced SLC20A1 in 690 individuals with bladder exstrophy-epispadias complex (BEEC) including 84 individuals with cloacal exstrophy. We identified two additional monoallelic de novo variants. One was identified in a case-parent trio with classic bladder exstrophy, and one additional novel de novo variant was detected in an affected mother who transmitted this variant to her affected son. To study the potential cellular impact of SLC20A1 variants, we expressed them in HEK293 cells. Here, phosphate transport was not compromised, suggesting that it is not a disease mechanism. However, there was a tendency for lower levels of cleaved caspase-3, perhaps implicating apoptosis pathways in the disease. Our results suggest SLC20A1 is involved in urinary tract and urorectal development and implicate SLC20A1 as a disease-gene for BEEC.
Barbosa Sicard, Eduardo ; Frömel, Timo ; Keserü, Benjamin ; Brandes, Ralf ; Morisseau, Christophe ; Hammock, Bruce D. ; Braun, Thomas ; Krüger, Marcus ; Fleming, Ingrid
Inhibition of the soluble epoxide hydrolase (sEH) has beneficial effects on vascular inflammation and hypertension indicating that the enzyme may be a promising target for drug development. As the enzymatic core of the hydrolase domain of the human sEH contains two tyrosine residues (Tyr383 and Tyr466) that are theoretically crucial for enzymatic activity, we addressed the hypothesis that the activity of the sEH may be affected by nitrosative stress. Epoxide hydrolase activity was detected in human and murine endothelial cells as well in HEK293 cells and could be inhibited by either authentic peroxynitrite (ONOO−) or the ONOO− generator 3-morpholino-sydnonimine (SIN-1). Protection of the enzymatic core with 1-adamantyl-3-cyclohexylurea in vitro decreased sensitivity to SIN-1. Both ONOO− and SIN-1 elicited the tyrosine nitration of the sEH protein and mass spectrometry analysis of tryptic fragments revealed nitration on several tyrosine residues including Tyr383 and Tyr466. Mutation of the latter residues to phenylalanine was sufficient to abrogate epoxide hydrolase activity. In vivo, streptozotocin-induced diabetes resulted in the tyrosine nitration of the sEH in murine lungs and a significant decrease in its activity. Taken together, these data indicate that the activity of the sEH can be regulated by the tyrosine nitration of the protein. Moreover, nitrosative stress would be expected to potentiate the physiological actions of arachidonic acid epoxides by preventing their metabolism to the corresponding diols.
Wiederstein, Janica Lea ; Nolte, Hendrik ; Günther, Stefan ; Piller, Tanja ; Baraldo, Martina ; Kostin, Sawa ; Bloch, Wilhelm ; Schindler, Natalie ; Sandri, Marco ; Blaauw, Bert ; Braun, Thomas ; Hölper, Soraya ; Krüger, Marcus
Protein aggregates and cytoplasmic vacuolization are major hallmarks of multisystem proteinopathies (MSPs) that lead to muscle weakness. Here, we identify METTL21C as a skeletal muscle-specific lysine methyltransferase. Insertion of a β-galactosidase cassette into the Mettl21c mouse locus revealed that METTL21C is specifically expressed in MYH7-positive skeletal muscle fibers. Ablation of the Mettl21c gene reduced endurance capacity and led to age-dependent accumulation of autophagic vacuoles in skeletal muscle. Denervation-induced muscle atrophy highlighted further impairments of autophagy-related proteins, including LC3, p62, and cathepsins, in Mettl21c−/− muscles. In addition, we demonstrate that METTL21C interacts with the ATPase p97 (VCP), which is mutated in various human MSP conditions. We reveal that METTL21C trimethylates p97 on the Lys315 residue and found that loss of this modification reduced p97 hexamer formation and ATPase activity in vivo. We conclude that the methyltransferase METTL21C is an important modulator of protein degradation in skeletal muscle under both normal and enhanced protein breakdown conditions.
Wiesmann, Frank ; Vachta, Jan ; Ehret, Robert ; Walter, Hauke ; Kaiser, Rolf ; Stürmer, Martin ; Tappe, André ; Däumer, Martin ; Berg, Thomas ; Naeth, Gudrun ; Braun, Patrick ; Knechten, Heribert
Background: Although being considered as a rarely observed HIV-1 protease mutation in clinical isolates, the L76V-prevalence increased 1998-2008 in some European countries most likely due to the approval of Lopinavir, Amprenavir and Darunavir which can select L76V. Beside an enhancement of resistance, L76V is also discussed to confer hypersusceptibility to the drugs Atazanavir and Saquinavir which might enable new treatment strategies by trying to take advantage of particular mutations. Results: Based on a cohort of 47 L76V-positive patients, we examined if there might exist a clinical advantage for L76V-positive patients concerning long-term success of PI-containing regimens in patients with limited therapy options. Genotypic- and phenotypic HIV-resistance tests from 47 mostly multi-resistant, L76V-positive patients throughout Germany were accomplished retrospectively 1999-2009. Five genotype-based drug-susceptibility predictions received from online interpretation-tools for Atazanavir, Saquinavir, Amprenavir and Lopinavir, were compared to phenotype-based predictions that were determined by using a recombinant virus assay along with a Virtual Phenotype™(Virco). The clinical outcome of the L76V-adapted follow-up therapy was determined by monitoring viral load for 96 weeks. Conclusions: In this analysis, the mostly used interpretation systems overestimated the L76V-mutation concerning Atazanavir- and SQV resistance. In fact, a clear benefit in drug susceptibility for these drugs was observed in phenotype analysis after establishment of L76V. More importantly, long-term therapy success was significantly higher in patients receiving Atazanavir and/or Saquinavir plus one L76V-selecting drug compared to patients without L76V-selecting agents (p = 0.002). In case of L76V-occurrence ATV and/or SQV may represent encouraging options for patients in deep salvage situations.
Obermeier, Martin ; Berg, Thomas ; Sichtig, Nadine ; Braun, Patrick ; Däumer, Martin P. ; Walter, Hauke ; Noah, Christian ; Wolf, Eva ; Müller, Harm ; Stürmer, Martin ; Thielen, Alexander ; Kaiser, Rolf
Poster presentation: Background Maraviroc is a new drug used to treat HIV infection from the new class of drugs called CCR5 entry inhibitors. As the active principle of these drugs is to block the CCR5-receptor on the surface of the target cells, it has to be known if the virus in the patient is using only CCR5 as co-receptor or if there are populations that can also use CXCR4. Therefore, an assay to determine the tropism has to be performed before starting a therapy. Besides phenotypic assays like the TROFILE® assay by Monogram, used in the approval studies, there exist several genotyping systems like geno2pheno-coreceptor, Wetcat (providing five different genotypic tropism schemes) and WebPSSM. ...
Prieto García, Cristian ; Hartmann, Oliver ; Reissland, Michaela ; Braun, Fabian ; Fischer, Thomas ; Walz, Susanne ; Schülein‐Völk, Christina ; Eilers, Ursula ; Ade, Carsten P. ; Calzado, Marco A. ; Orian, Amir ; Maric, Hans-Michael ; Münch, Christian ; Rosenfeldt, Mathias Tillmann ; Eilers, Martin ; Diefenbacher, Markus Elmar
The transcription factor ∆Np63 is a master regulator of epithelial cell identity and essential for the survival of squamous cell carcinoma (SCC) of lung, head and neck, oesophagus, cervix and skin. Here, we report that the deubiquitylase USP28 stabilizes ∆Np63 and maintains elevated ∆NP63 levels in SCC by counteracting its proteasome‐mediated degradation. Impaired USP28 activity, either genetically or pharmacologically, abrogates the transcriptional identity and suppresses growth and survival of human SCC cells. CRISPR/Cas9‐engineered in vivo mouse models establish that endogenous USP28 is strictly required for both induction and maintenance of lung SCC. Our data strongly suggest that targeting ∆Np63 abundance via inhibition of USP28 is a promising strategy for the treatment of SCC tumours.
Bobe, René ; Aldeias, Vera ; Alemseged, Zeresenay ; Archer, Will ; Aumaître, Georges ; Bamford, Marion K. ; Biro, Dora ; Bourlès, Didier L. ; Braun, David R. ; Capelli, Cristian ; d’Oliveira Coelho, João ; Habermann, Jörg M. ; Head, Jason J. ; Keddadouche, Karim ; Kupczik, Kornelius ; Lebatard, Anne-Elisabeth ; Lüdecke, Tina ; Macôa, Amélia ; Martı́nez, Felipe I. ; Mathe, Jacinto ; Mendes, Clara ; Meira Paulo, Luis ; Pinto, Maria ; Püschel, Thomas A. ; Regala, Frederico Tátá ; Sier, Mark Jan ; da Silva, Maria Joana Ferreira ; Stalmans, Marc ; Carvalho, Susana
The Miocene is a key time in the evolution of African mammals and their ecosystems witnessing the origin of the African apes and the isolation of eastern coastal forests through an expanding biogeographic arid corridor. Until recently, however, Miocene sites from the southeastern regions of the continent were unknown. Here we report discovery of the first Miocene fossil teeth from the shoulders of the Urema Rift in Gorongosa National Park, Mozambique, at the southern East African Rift System. We provide the first 1) radiometric age determinations of the fossiliferous Mazamba Formation, 2) reconstructions of past vegetation in the region based on pedogenic carbonates and fossil wood, and 3) description of fossil teeth from the southern rift. Gorongosa is unique in the East African Rift System in combining marine invertebrates, marine vertebrates, terrestrial mammals, and fossil woods in coastal paleoenvironments. The Gorongosa fossil sites offer the first evidence of persistent woodlands and forests on the coastal margins of southeastern Africa during the Miocene, and an exceptional assemblage of fossil vertebrates including new species. Further work will allow the testing of hypotheses positing the formation of a northeast-southwest arid corridor isolating species on the eastern coastal forests from those elsewhere in Africa.
Brief The Miocene is a key time in the evolution of African mammals and their ecosystems encompassing hominine origins and the establishment of an arid corridor that isolated eastern Africa’s coastal forests. Until now, however, Miocene sites from southeastern Africa have been unknown. We report the discovery of the first Miocene fossil sites from Gorongosa National Park, Mozambique, and show that these sites formed in coastal settings. We provide radiometric ages for the fossiliferous sediments, reconstructions of past vegetation based on stable isotopes and fossil wood, and a description of the first fossil teeth from the region. Gorongosa is the only paleontological site in the East African Rift that combines fossil woods, marine invertebrates, marine vertebrates, and terrestrial mammals. Gorongosa offers the first evidence of persistent woodlands and forests on the coastal margins of southeastern Africa during the Miocene.
Bobe, René ; Aldeias, Vera ; Alemseged, Zeresenay ; Anemone, Robert L. ; Archer, Will ; Aumaître, Georges ; Bamford, Marion K. ; Biro, Dora ; Bourlès, Didier L. ; Boyd, Melissa Doyle ; Braun, David R. ; Capelli, Cristian ; Coelho, João d’Oliveira ; Habermann, Jörg M. ; Head, Jason J. ; Keddadouche, Karim ; Kupczik, Kornelius ; Lebatard, Anne-Elisabeth ; Lüdecke, Tina ; Macôa, Amélia ; Martínez, Felipe I. ; Mathe, Jacinto ; Mendes, Clara ; Meira Paulo, Luis ; Pinto, Maria ; Presnyakova, Darya ; Püschel, Thomas A. ; Regala, Frederico Tátá ; Sier, Mark Jan ; Silva, Maria Joana Ferreira da Silva ; Stalmans, Marc ; Carvalho, Susana
The Miocene was a key time in the evolution of African ecosystems witnessing the origin of the African apes and the isolation of eastern coastal forests through an expanding arid corridor. Until recently, however, Miocene sites from the southeastern regions of the continent were unknown. Here, we report the first Miocene fossil teeth from the shoulders of the Urema Rift in Gorongosa National Park, Mozambique. We provide the first 1) radiometric ages of the Mazamba Formation, 2) reconstructions of paleovegetation in the region based on pedogenic carbonates and fossil wood, and 3) descriptions of fossil teeth. Gorongosa is unique in the East African Rift in combining marine invertebrates, marine vertebrates, reptiles, terrestrial mammals, and fossil woods in coastal paleoenvironments. The Gorongosa fossil sites offer the first evidence of woodlands and forests on the coastal margins of southeastern Africa during the Miocene, and an exceptional assemblage of fossils including new species.
Die Zytomegafie ist eine meist lebenslänglich latent bleibende vertikale und horizontale Herpesvirusinfektion mit gelegentlich schweren Krankheitsbildern, auch als Ursache oder Folge von Immunstörungen. Dem Virus wird ein onkogenes Potential zugeschrieben, zuletzt diskutiert bei AIDS und M. Kaposi. Für die Labordiagnose verfügen wir über die Mikroskopie (Zellkerneinschlüsse) und Elektronenmikroskopie, Nachweis der Virusinfektiosität auf Zellkulturen, DNA- und Polypeptidanalyse zur Virusstammidentifikation, direkte DNA- und Antigennachweise aus Patientenmaterial, immunhistologische Methoden (z.B. Immunperoxydase- Technik). Die Untersuchung der Immunzellen erfolgt bei der Zytomegalie quantitativ (T-ZellQuotient) und qualitativ (Lymphozytenstimulierung, neuerdings auch mit Vollblut). Am leichtesten gelingt die Labordiagnose serologisch, d.h. über den Antikörpernachweis. Dafür sind eine Vielzahl „liquid" und „solid phase"-Assays entwickelt worden. Am meisten haben sich heute neben der KB R (und P H A) Immunofluororeszenz und ELISA durchgesetzt, wobei einerseits unterschiedliche Antigene („early", „late antigens") und Antigenpräparationen (z. B. Viruskapsid, -envelope) zum Einsatz kommen, andererseits verschiedene Ig- Klassen und -Subklassen getestet werden, um die primäre und sekundäre Zytomegalie zu diagnostizieren und zu differenzieren. Speziell für den Ig M -Nachweis wurden viele Testmodifikationen etabliert; Rheumafaktorinterferenz und IgG-Kompetition lassen sich am besten durch IgG-Präzipitation ausschalten. Die neuen Methoden haben nicht nur die Aufklärung vieler interessanter Krankheitsfälle, sondern auch exakte epidemiologische Studien bei Risikogruppen ermöglicht (Blutspender: 47[0], schwangere Frauen 56[13], Patienten mit Hämophilie: 69[0], nach NTPL: 90[24], nach Herz-OP: 87[1], Prostituierte: 90[1]% CMV-IgG[(IgM)- Antikörperträger].