CT-guided lung biopsy has a high diagnostic accuracy. Complication rates were acceptable and comparable to those of previous studies.

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BackgroundThe aim of this study was to detect the epidermal growth factor receptor (EGFR)-activating mutations and other oncogene alterations in patients with non-small-cell lung cancers (NSCLC) who experienced a treatment failure in response to EGFR-tyrosine kinase inhibitors (TKIs) with a next generation sequencer.MethodsFifteen patients with advanced NSCLC previously treated with EGFR-TKIs were examined between August 2005 and October 2014. For each case, new biopsies were performed, followed by DNA sequencing on an Ion Torrent Personal Genome Machine (PGM) system using the Ion AmpliSeq Cancer Hotspot Panel version 2.ResultsAll 15 patients were diagnosed with NSCLC harboring EGFR-activating mutations (seven cases of exon 19 deletion, seven cases of L858R in exon 21, and one case of L861Q in exon 21). Of the 15 cases, acquired T790M resistance mutations were detected in 9 (60.0 %) patients. In addition, other mutations were identified outside of EGFR, including 13 cases (86.7 %) exhibiting TP53 P72R mutations, 5 cases (33.3 %) of KDR Q472H, and 2 cases (13.3 %) of KIT M541L.ConclusionsHere, we showed that next-generation sequencing (NGS) is able to detect EGFR T790M mutations in cases not readily diagnosed by other conventional methods. Significant differences in the degree of EGFR T790M and other EGFR-activating mutations may be indicative of the heterogeneity of disease phenotype evident within these patients. The co-existence of known oncogenic mutations within each of these patients may play a role in acquired EGFR-TKIs resistance, suggesting the need for alternative treatment strategies, with PCR-based NGS playing an important role in disease diagnosis.

We have used fragments of the TSH receptor (TSHR) expressed in E. coli as glutathione S-transferase fusion proteins to produce rabbit polyclonal antibodies and a panel (n=5) of monoclonal antibodies to the extracellular fragment of the TSHR. The binding characteristics of the antibodies to linear, conformational, glycosylated and unglycosylated forms of the receptor in different assay systems have been investigated.The reactivity of these antibodies with the TSHR was assessed by Western blotting with both native and recombinant human TSHR expressed in CHO cells, immunoprecipitation of 35 S-labelled fulllength TSHR produced in an in vitro transcription/ translation system, immunoprecipitation of 125 I-TSH/TSHR complexes, inhibition of 125 I-TSH binding to the TSHR and fluorescence activated cell sorter (FACS) analysis of binding to CHO-K1 cells expressing the TSHR on their cell surface. Fab fragments of monoclonal antibodies were isolated, labelled with 125 I and used to determine the affinity constants of these antibodies with receptor, bound and free Fab being separated by polyethylene glycol (PEG) precipitation.Rabbit polyclonal and mouse monoclonal antibodies reacted with the TSHR in Western blotting and one monoclonal antibody (3C7) was able to inhibit 125 I-TSH binding to native human TSHR (74% inhibition), recombinant human TSHR (84% inhibition) and porcine TSHR (65% inhibition). Affinity constant values for TSHR monoclonal antibody Fab fragments calculated using Scatchard analysis were about 10 7 M 1 . Four out of five monoclonal antibodies reacted in FACS analysis with TSHR expressed on the surface of CHO-K1 cells. The FACS unreactive monoclonal (3C7) bound well to detergent solubilised TSH receptors and this emphasised the importance of using a combination of FACS analysis and radioactivelylabelled probes in analysis of the TSH receptor.The monoclonal antibodies produced in this study were found to be of relatively low affinity but proved useful for detection of the receptor by Western blotting and by FACS analysis.

Our data indicate that the personalized 3D printed model of the kidney is useful for preoperative education among patients and their families who underwent RAPN, especially in elderly people.