• The workflow is depicted in the following image:

• In the first step, the bacterial cells are concentrated by filtration (this step is also part of the standard method). The water sample is filtered with a standard filtration unit via a polycarbonate filter with a pore size of 0.22 μm, supplied in the rqmicro kits. The filter is then resuspended in a volume of 3 mL buffer.
• Specific antibodies bound to magnetic particles as well as specific antibodies bound to a fluorescent dye are then added to the sample. The antibodies bound to the magnetic particles enable isolation of the target cells from the rest of the sample by means of the so-called immunomagnetic separation (IMS). The antibodies bound to a fluorescent dye allow the quantification of the bacterial cells after the immunomagnetic separation in a flow cytometer.
• The sample is then incubated for 30 minutes at room temperature so that the antibodies can bind to the target cells.
• During the separation process, which takes place either in the rqmicro CellStream instrument in a microfluidic cartridge or in the Manual Immunomagnetic Separator (MIMS), the bacterial target cells (for example, Legionella) are collected in a volume of 1 mL of clean buffer (second concentration step). With the rqmicro sample preparation a sample purity of more than 95% is achieved.
• The positive fraction can then be measured and quantified using a commercially available flow cytometer. However, the sample can also be analyzed by PCR, by plating and culturing of bacterial cells (standard method), fluorescence microscopy, etc.

• During the separation process, which takes place in the CellStream instrument in a microfluidic cartridge, the sample is transported through a narrow channel from left to right. A vacuum pump adds positive or negative pressure in the corresponding containers of the cartridge and thereby creates this flux. During this process, a magnet is lowered down onto the microfluidic channels and thereby prevents the bacterial cells, which are bound to magnetic particles via specific antibodies, from continuing to flow in the channel (immobilization). Thus, the target cells are immobilized and at the same time > 95% of the non-target cells (competing flora) originally present in the sample reach the residual fraction (Waste). This is followed by a wash step with buffer, which contributes to the high sample purity.
• In the last step of the separation, the magnet is raised and as a result, the concentrated and purified target cells are eluted in 1 mL of clean buffer into the container of the positive fraction.

• The immunomagnetic separation (IMS) takes place on a microfluidic cartridge in the CellStream instrument.

• The workflow is significantly shorter (time savings)
• It is possible to differentiate between viable and dead cells
• Enrichment is unnecessary
• Risk of false-negative results is significantly lower

Read full FAQ