Diagnostics

Bilder fehlen noch…

The following section summarizes the most interesting results we obtained this summer. All together we established a cell-free expression system that can be used to translate proteins from DNA immobilized on a PDMS (gibt es ein Glossar dafür?)slide. Proteins were expressed with a specific tag allowing its binding to the specific surface on the opposite slide. (The expressed protein is then attached to another slide via a specific tag system resulting in a distinct pattern.) The setup of the DiaCHIP based on a microfluidic system and antibody solutions can be flushed over the protein slide. (A microfluidic system is used to flush the slide with an antibody solution.) Specific antibody-antigen interactions were successfully detected with the label-free and real-time technology iRIf (imaging Reflectometric Interference). For diagnostic applications, the immobilized proteins are antigenic peptides specific for a certain pathogen.

BÄÄÄM1 - Salmonella

We obtained the sequence for a specific Salmonella Typhimurium antigen and a corresponding antibody from the lab of Prof. Dr. Hust. We successfully expressed both proteins in E. coli and spotted the His-tagged antigen on a specific PDITC surface. With the following iRIf measurement we (wanted to) analyzed the binding between antigen and antibody using our own DiaCHIP. The measurement was a great success, showing a distinct shift in the binding curve (habt ihr tatsächlich eine Bindekurve mit eurem Device erhalten?) for the salmonella antigen when the salmonella antibody is flushed over the chip, whereas the negative control is not bound by the antibody. With this measurement a proof of concept can be provided, showing that our device is indeed capable of detecting specific antigen-antibody binding, as it will be occurring when patient samples are analyzed.

BÄMMM2 – Tetanus

was ist der unterschied zu dem Inhalt auf der Result Summary Seite? Sollte hier nicht etwas mehr detaillierter stehen?

To show that our DiaCHIP does not only work with pure antibody solutions, but can detect antibodies in “real-life” serum samples, we did the following experiment with tetanus antigens and antibodies. After cloning the sequence coding for a specific tetanus antigen in an appropriate expression vector we expressed sufficient amounts of the antigen to spot it on a specific Ni-NTA surface. We obtained serum samples of a patient before and after tetanus vaccination that were flushed over the immobilized tetanus antigens using our DiaCHIP setup. With this experiment we additionally (intend to) show that we can distinguish between different antibody titers in a patient’s blood sample. We were indeed able to show a significant difference in the “before” and “after” sample (Fig 1 and Fig. 2). Thus, we could show that our DiaCHIP is a suitable device to analyze blood samples and not only detect specific antibodies against certain diseases but also differentiate between antibody levels.

BÄÄÄMM3 – Own device

In collaboration with the AG Roth from the ZBSA Freiburg we were allowed to use their iRIf (imaging reflectometric interference) setup to perform our measurements (with the DiaCHIP). We soon started to further investigate the physical mechanism of a device needed for our applications and planned to build our very own measuring device. After weeks of calculation and lots of try-and-error approaches we can now present our own DiaCHIP measuring device. The complete setup can be seen in Fig. X, where the different components are indicated.