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Mittenwalde, Germany

Negri P.,University of Georgia | Chen G.,University of Georgia | Kage A.,Aptares AG | Nitsche A.,Robert Koch Institute | And 3 more authors.
Analytical Chemistry

We have demonstrated label-free optical detection of viral nucleoprotein binding to a polyvalent anti-influenza aptamer by monitoring the surface-enhanced Raman (SERS) spectra of the aptamer-nucleoprotein complex. The SERS spectra demonstrated that selective binding of the aptamer-nucleoprotein complex could be differentiated from that of the aptamer alone based solely on the direct spectral signature for the aptamer-nucleoprotein complex. Multivariate statistical methods, including principal components analysis, hierarchical clustering, and partial least squares, were used to confirm statistically significant differences between the spectra of the aptamer-nucleoprotein complex and the spectra of the unbound aptamer. Two separate negative controls were used to evaluate the specificity of binding of the viral nucleoproteins to this aptamer. In both cases, no spectral changes were observed that showed protein binding to the control surfaces, indicating a high degree of specificity for the binding of influenza viral nucleoproteins only to the influenza-specific aptamer. Statistical analysis of the spectra supports this interpretation. AFM images demonstrate morphological changes consistent with formation of the influenza aptamer-nucleoprotein complex. These results provide the first evidence for the use of aptamer-modified SERS substrates as diagnostic tools for influenza virus detection in a complex biological matrix. © 2012 American Chemical Society. Source

Negri P.,University of Georgia | Kage A.,Aptares AG | Nitsche A.,Robert Koch Institute | Naumann D.,Robert Koch Institute | Dluhy R.A.,University of Georgia
Chemical Communications

A highly sensitive surface-enhanced Raman (SERS)-based method for detection of influenza viral nucleoproteins is described. The intrinsic SERS spectrum of the aptamer-nucleoprotein complex provides direct evidence of binding between a polyvalent anti-influenza aptamer and the nucleoproteins of three influenza strains. © The Royal Society of Chemistry 2011. Source

The invention relates to a method of selection, by two-dimensional separation, of nucleic acids that bind to a target molecule with high affinity from a mixture of nucleic acids, including the following steps: a) subjecting the mixture of nucleic acids to a physico-chemical separation step, thereby obtaining a set of mixed fractions containing the nucleic acids, a run parameter window being associated with every mixed fraction containing the nucleic acids, b) contacting a mixed fraction containing the nucleic acids with the target molecule, thereby obtaining a binding mixture containing nucleic acid/target molecule complexes, c) subjecting the binding mixture from step b) to the same physico-chemical separation step as in step a), thereby selecting nucleic acid/target molecule complexes whose run parameters are outside of the run parameter window.

The invention relates to a method for selecting nucleic acids that bond with high affinity to a target molecule from a mixture of nucleic acids, comprising: a) loading a column with the target molecules, b) feeding the nucleic acids into a first end of the column, to create a defined volumetric flow of medium through the column, c) immobilizing the nucleic acids to the target molecule wherein an affinity of the nucleic acids to the target molecule decreases as the distance from the first end of the column increases, d) stopping the volumetric flow after a defined period of time, e) cutting the column into segments, and allocating a routing co-ordinate to each segment, and f) identifying and collecting nucleic acids that bond with a high affinity to the target molecule by desorbing the immobilized nucleic acids from at least one segment.

Wallukat G.,Max Delbruck Centrum fur Molekulare Medizin | Wallukat G.,Aptares AG | Haberland A.,Charite - Medical University of Berlin | Berg S.,Institute For Diabetes Gerhardt Katsch | And 9 more authors.
Circulation Journal

Background: Application of immunoapheresis to eliminate pathogenic autoantibodies targeting the second extracellular loop of the β1-receptor (β1-AABs) is currently investigated in patients with cardiomyopathy. Aptamers (single short DNA or RNA strands) are a new class of molecules that bind to a specific target molecule. This property qualifies aptamers for potential use in the apheresis technique. We recently identified an aptamer that specifically binds to β1-AABs, so in the present study we tested whether this aptamer could be used as a binder to prepare an apheresis column suitable for clearing β1-AABs from rat's blood. Methods and Results: An apheresis column was designed containing the β1-AAB-targeting-aptamer coupled to sepharose. As tested in vitro, this column (1) binds β1-AABs highly specifically without marked interference with common IgGs, (2) has a capacity for clearing of approximately 1L of β1-AAB-positive serum and (3) can be completely regenerated for subsequent use. Using the column for extracorporeal apheresis of spontaneously hypertensive rats (SHR) positive for both β1-AABs and muscarinic 2-receptor autoantibodies (M2-AABs), only β1-AABs were removed. In a follow-up of 9 weeks, recurrence of β1-AABs in the blood of SHR could not be detected. Conclusions: For the first time, a newly designed apheresis column with a β1-AAB specific aptamer as a binder was successfully used to eliminate β1-AABs from SHR blood. Source

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