Angelika Lueking

Protagen AG · Diagnostics

Objective: To assess the diagnostic potential of IgG antibodies to citrullinated and corresponding native autoantigens in early arthritis. Methods: IgG autoantibodies to 390 distinct unmodified and corresponding in vitro citrullinated recombinant proteins were measured by a multiplex assay in baseline blood samples from a German multicenter nati...

The classification of a population by a specific trait is a major task in medicine, for example when in a diagnostic setting groups of patients with specific diseases are identified, but also when in predictive medicine a group of patients is classified into specific disease severity classes that might profit from different treatments. When the siz...

Background The aim was to identify novel diagnostic autoantibody candidates for rheumatoid arthritis (RA) by comprehensive screening for autoreactivity. Method We incubated 5892 recombinant proteins coupled to fluorescent beads, with patients’ sera for the detection of IgG-autoantibodies in three independent patient cohorts: A (n = 72 patients with...

Background High throughput protein expression studies can be performed using bead-based protein immunoassays, such as the Luminex® xMAP® technology. Technical variability is inherent to these experiments and may lead to systematic bias and reduced power. To reduce technical variability, data pre-processing is performed. However, no recommendations...

Systemic lupus erythematosus (SLE) is a heterogeneous disease with respect to disease manifestations, disease progression and treatment response. Therefore, strategies to identify biomarkers that help distinguishing SLE subgroups are a major focus of biomarker research. We reasoned that a multiparametric autoantibody profiling approach combined wit...

Background: Chronic inflammation is frequently observed on histological analysis of malignant and non-malignant prostate specimens. It is a suspected supporting factor for prostate diseases and their progression and a main cause of false positive PSA tests in cancer screening. We hypothesized that inflammation induces autoantibodies, which may be...

Clinical parameters are equally distributed between high and low inflammation patient cohorts. All samples were tested for statistically significant differences between the two sample groups. Clinical parameters including age, C-reactive protein, Gleason Score, prostate volume, PSA and free PSA (Table 1) are equally distributed amongst the high and...

Autoantibody marker antigens are differentially expressed in benign tissue and malignant lesions of the prostate. A Representative images for SPAST, STX18 and SPOP immunohistochemical double stainings with the basal cell marker p63. Left side shows benign lesions, right side represents invasive prostate carcinoma. The basal cell marker, which is ab...

List of autoantigens included in the planar protein microarray (Table "Protein Array") and in the Luminex bead protein array (Table "Luminex"). Autoantigen proteins included in the planar protein array or in the Luminex bead protein array are listed. The number of positive samples of indivual auoantigens for the low and the high inflammation cohort...

List of 165 significantly upregulated autoantibodies in serum samples of the screening cohort, ranked according to p-Value. (XLSX)

Background The discovery of autoantibodies against citrullinated proteins (ACPA) and the consequent development of serological tests using cyclic citrullinated peptide (CCP) have improved diagnosis of rheumatoid arthritis (RA). ACPAs recognize specifically various citrullinated peptides that derive from different antigens. It has also been demonstr...

We have previously proposed that specific proteins may form insoluble aggregates as a response to an illness-specific proteostatic dysbalance in a subset of brains from individuals with mental illness, as is the case for other chronic brain conditions. So far, established risk factors DISC1 and dysbindin were seen to specifically aggregate in a sub...

Background SLE is a chronic multisystem autoimmune disease with yet unknown etiology. One hallmark of SLE is the B cell activation and the production of various autoantibodies. Differential diagnosis involves ANA screening which may yield also positive results in many connective tissue disorders and other autoimmune diseases, and may occur in norma...

Background Rheumatoid arthritis (RA) is an autoimmune disease typically characterized by chronic inflammation, accumulation of self-reactive B-cells and production of autoantibodies of which anti–cyclic citrullinated peptide (anti-CCP) antibodies and rheumatoid factor (RF) have diagnostic utility. Despite these two well established markers up to 30...

Huntington's disease (HD) is a progressive neurodegenerative disorder, which is caused by expansion of a polyglutamine tract in the first exon of the huntingtin gene. Genetic testing allows the unambiguous diagnose of HD and the identification of individuals carrying the gene defect before onset of clinical symptoms. However, molecular markers that...

The stability of therapeutic antibodies during downstream processing and storage is important for functionality and quality. To determine functional antibody performance, the UNIchip® high-density protein microarray with 384 recombinant antigenic targets was developed; this allows characterization of antibody specificity by generating standardized...

The currently used prostate cancer serum marker has a low cancer specificity and improved diagnostics are needed. Here we evaluated whether autoantibodies are present in sera of prostate cancer patients and whether they are useful diagnostic markers for prostate cancer. Sera from 20 prostate cancer patients and 20 healthy controls were incubated on...

Using the UNIarray® marker technology platform, cerebrospinal fluid immunoglobulin G reactivities of 15 controls and 17 RRMS patients against human recombinant proteins were investigated. Patient cerebrospinal fluids were oligoclonal band positive and reactivities were compared to that of sex- and age-matched controls. We hereby aimed at the charac...

Validation of the VZV RecomLine with clinically defined serum samples. This table depicts all serological parameters of the clinically defined patient sera.VZV-IgG and IgM ELISA titres were assayed by wcELISA (Dade Behring, Enzygnost, Germany). Qualitative RecomLine VZV IgG and IgM reactivities towards individual antigens are depicted as "1" (react...

Analysis of possible crossreactivities of the RecomLine VZV recombinant antigens with serum samples of defined antibody status to HSV. This table depicts the cross-reactivity of all serologically defined VZV-IgG negative patient samples according to their HSV (IgG) status. No cross-reactivity in the in the RecomLine VZV IgM assay can be observed. T...

Varicella-Zoster virus causes chickenpox upon primary infection and shingles after reactivation. Currently available serological tests to detect VZV-specific antibodies are exclusively based on antigens derived from VZV-infected cells. We present a systematic approach for the identification of novel, serologically reactive VZV antigens. Therefore,...

We describe a novel procedure for determination and validation of off-target activities of anti-cluster designation antigen identity 44 (CD44) variant 6 recombinant antibodies by combining two complementary technology platforms; namely UNIchip AV-400, containing a printed serial dilution of CD44 variant 6 and approximately 400 different human prote...

Tumor necrosis factor-alpha inhibitors are widely and successfully used to treat rheumatic diseases. However, significant side effects have been reported. To detect the potential off-target activities of such inhibitors we characterized two therapeutic antibodies (adalimumab, infliximab) and one receptor fusion protein (etanercept) on protein bioch...

Protein Microarrays enable the analysis of multiple binding events in parallel. Based on patented UNIclone technology, we have established a reproducible protein biochip platform to determine specific binding profiles to multiple targets in a quantitative way. The UNIchip protein biochips provide a novel and unique tool for antibody characterizatio...

High density DNA microarray technology has played a key role in the analysis of whole genomes and their gene expression patterns. The ability to study many thousands of individual genes using oligonucleotide or cDNA arrays is now very widespread, with its uses ranging from the profiling of gene expression patterns in whole organisms or tissues to t...

Protein microarrays with immobilised proteins on their surface are new analytical tools to overcome the current limits with respect to sample volume and throughput. They have a great potential as well with respect to multiplexing of complex samples, as a research tool and in diagnostics. Based on recent advances in this technology, new applications...

Dilated cardiomyopathy (DCM) is a myocardial disease characterized by progressive depression of myocardial contractile function and ventricular dilatation. Thirty percent of DCM patients belong to the inherited genetic form; the rest may be idiopathic, viral, autoimmune, or immune-mediated associated with a viral infection. Disturbances in humoral...

Array technology enables genome-wide screening for protein expression and interactions at high throughput. Protein microarrays are emerging as a major technology platform for functional genomics.

Protein biochips have a great potential in future parallel processing of complex samples as a research tool and in diagnostics. For the generation of protein biochips, highly automated technologies have been developed for cDNA expression library production, high throughput protein expression, large scale analysis of proteins, and protein microarray...

The human genome has been sequenced and the challenges of understanding the function of the newly discovered genes have been addressed. High-throughput technologies such as DNA microarrays have been developed for the profiling of gene expression patterns in whole organisms or tissues. Protein arrays are emerging to follow DNA chips as possible scre...

The mouse is the premier genetic model organism for the study of disease and development. We describe the establishment of a mouse T helper cell type 1 (T(H)1) protein expression library that provides direct access to thousands of recombinant mouse proteins, in particular those associated with immune responses. The advantage of a system based on th...

The expression and characterization of large protein libraries requires high-throughput tools for rapid and cost-effective expression and screening. A promising tool to meet these requirements is miniaturized high-density plates in chip format, consisting of an array of wells with submicroliter volumes. Here, we show the combination of nanowell chi...

There is burgeoning interest in protein microarrays, but a source of thousands of nonredundant, purified proteins was not previously available. Here we show a glass chip containing 2413 nonredundant purified human fusion proteins on a polymer surface, where densities up to 1600 proteins/cm(2) on a microscope slide can be realized. In addition, the...

Protein array technology has emerged as a new tool to enable ordered screening of proteins for expression and molecular interactions in high throughput. Besides classical solid-phase substrates, such as micro-titre plates and membrane filters, protein arrays have recently been devised with chip-sized supports. Several applications on protein chips...

High-throughput protein arrays allow the miniaturized and parallel analysis of large numbers of diagnostic markers in complex samples. Using automated colony picking and gridding, cDNA or antibody libraries can be expressed and screened as clone arrays. Protein microarrays are constructed from recombinantly expressed, purified, and yet functional p...

The generation of protein chips requires much more efforts than DNA microchips. While DNA is DNA and a variety of different DNA molecules behave stable in a hybridisation experiment, proteins are much more difficult to produce and to handle. Outside of a narrow range of environmental conditions, proteins will denature, lose their three-dimensional...

We describe the technical feasibility and methodology to characterize a protein by a minimal set of structural information generated by matrix assisted laser desorption/ionization (MALDI)-mass spectrometry, termed a "minimal protein Identifier" (MPI). MPIs can be determined for proteins from two-dimensional gels and recombinant proteins and can be...

Large-scale and high throughput approaches increasingly play an essential role in the study of biological systems, which are per se highly complex. Therefore, they need to be examined by these extensive methods to receive information about the large genomic and proteomic networks. In plant biology, this purpose has a strong support through the acce...

DNA-chip analysis has come a long way since the first conference in Moscow in 1991. Nowadays, DNA-microarrays seem to be a common commodity in biological sciences. The complexity hidden behind the apparent ease of such studies, however, is highlighted by the fact that it took about ten years before the methodology really set off. Also, on closer sc...

We developed a high-throughput technique for the generation of cDNA libraries in the yeast Saccharomyces cerevisiae which enables the selection of cloned cDNA inserts containing open reading frames (ORFs). For direct screening of random-primed cDNA libraries, we have constructed a yeast shuttle/expression vector, the so-called ORF vector pYEXTSH3,...

The human genome is sequenced and the challenges of understanding the function of the newly discovered genes have been addressed. For this purpose, high-throughput technologies have been developed that allow the monitoring of gene activity at the transcriptional level by analysis of complex expression patterns of a specific tissue. Differential gen...

The human genome is sequenced, but only a minority of genes have been assigned a function. Whole-genome expression profiling is an important tool for functional genomic studies. Automated technology allows high-throughput gene activity monitoring by analysis of complex expression patterns, resulting in fingerprints of diseased versus normal or deve...

We have constructed a novel Pichia pastoris/Escherichia coli dual expression vector for the production of recombinant proteins in both host systems. In this vector, an E. coli T7 promoter region, including the ribosome binding site from the phage T7 major capsid protein for efficient translation is placed downstream from the yeast alcohol oxidase p...

The array format has revolutionised biomedical experimentation and diagnostics, enabling ordered high-throughput analysis. During the past decade, classic solid phase substrates, such as microtitre plates, membrane filters and microscopic slides, were turned into high-density, chip-like structures. The concept of the arrayed library was central to...

Proteins translate genomic sequence information into function, enabling biological processes. As a complementary approach to gene expression profiling on cDNA microarrays, we have developed a technique for high-throughput gene expression and antibody screening on chip-size protein microarrays. Using a picking/spotting robot equipped with a new tran...

RNA helicases represent a large family of proteins that have been detected in almost all biological systems where RNA plays a central role. They are ubiquitously distributed over a wide range of organisms and are involved in nuclear and mitochondrial splicing processes, RNA editing, rRNA processing, translation initiation, nuclear mRNA export, and...