FAS2FURIOUS: Moderate-Throughput Secreted Expression of Difficult Recombinant Proteins in Drosophila S2 Cells.

Recombinant protein expression in eukaryotic insect cells is a powerful approach for producing challenging targets. However, due to incompatibility with standard baculoviral platforms and existing low-throughput methodology, the use of the Drosophila melanogaster "S2" cell line lags behind more common insect cell lines such as Sf9 or High-Five™. Due to the advantages of S2 cells, particularly for secreted and secretable proteins, the lack of a simple and parallelizable S2-based platform represents a bottleneck, particularly for biochemical and biophysical laboratories. Therefore, we developed FAS2FURIOUS, a simple and rapid S2 expression pipeline built upon an existing low-throughput commercial platform. FAS2FURIOUS is comparable in effort to simple E. coli systems and allows users to clone and test up to 46 constructs in just 2 weeks. Given the ability of S2 cells to express challenging targets, including receptor ectodomains, secreted glycoproteins, and viral antigens, FAS2FURIOUS represents an attractive orthogonal approach for protein expression in eukaryotic cells.

[An epidural hematoma following single shot epidural anesthesia]. / Epidurales Hämatom nach Single shot-Epiduralanästhesie.

The formation of an epidural or subdural haematoma is a well-known but very rare complication when anaesthetic procedures are conducted near the spinal cord. The major reasons are impaired blood coagulation and the trauma of puncture, which represents the initiating factor and determines the extent of the vascular lesion, while defective coagulation may cause the bleeding to continue. We report on a 30-year-old slender female patient of ASA group II undergoing epidural anaesthesia at L3/4 with a 19-gauge Crawford needle for an ankle joint injury. Unexpected puncture difficulties made several approaches necessary. Prevention of thrombosis with low-molecular heparin was not started preoperatively but 8 h following the intervention. On the first postoperative day the patient was mobilized and, in spite of the repeated punctures, there were no complaints until the fourth postoperative day. Subsequently, sudden significant complaints occurred, first suggesting infection near the spinal cord. Immediate magnetic resonance imaging revealed an epidural haematoma, which did not require intervention. Puncture of the spinal fluid was not performed. Laboratory analysis of coagulation showed a prothrombin time of 56%; PTT was normal with 36 s. Following three days of symptomatic therapy, the complaints had improved markedly, so that mobilization could be started again. For another 10 days the patient had a minor diffuse sensation of pressure when moving the lumbar spine in this region. Subsequently, there were no complaints at all, as proved by an investigation one year later. Unfortunately, the patient refused to undergo another MRT at that time. In the present case the haematoma was caused by tissue trauma due to several puncture attempts.(ABSTRACT TRUNCATED AT 250 WORDS)