To harvest human keratinocyte growth factor (KGF) mimic peptides with Ph.D.-7TM phage display peptide l ibrary. Methods Ph.D.-7TM phage display peptide l ibrary was biopanned for 4 rounds to harvest monoclonal anti-body human KGF and then phage tilter was detected. ELISA detection was performed to detect the binding force of random-selected monoclonal phages, thereafter DNA extracted from phages with better binding activity was sequenced and the Basic BLAST system was appl ied to conduct the sequence similarity and homology analysis. Results After 4 rounds ofbiopanning, the titer of phages was increased gradually and the enrichment of specific phage mimic peptides was obtained. The titers of monoclonal phages were up to 2.0 × 1014 pfu/mL according to ELISA detection. According to the absorbance value, the monoclonal phages with better binding activities to certain specific antibodies were sequenced, and 26 base sequences related to the promotion of division and growth were verified, 2 of which were similar to human KGF. Homology sequence analysis revealed that the common sequence of those 26 base sequences was similar to the partial sequences of human KGF. Conclusion The phage mimic peptides resembl ing or related to human KGF DNA can be harvested from Ph.D.-7TM phage display peptide l ibrary, which may be conducive to improve human KGF performance, wound heal ing and the qual ity of tissue engineered skin substitutes.
【Abstract】ObjectiveTo develop a new singletube polymerase chain reaction amplification (ST Amp) protocol for the efficient sequencing-based typing (SBT) of human leukocyte antigen DRB1（HLA-DRB1）.MethodsA set of 7 group-specific exonic 5′ amplification primers and a single generic 3′ primer were included together in a single PCR mix to facilitate a single PCR amplification per sample for HLA-DRB1 typing.ResultsAll samples were successfully typed, the typing result was accurate and repeatable.ConclusionST Amp technique has resulted in the ability to perform high-resolution, high-specificity and high-throughput HLA-DRB1 typing by DNA sequencing.
ObjectiveTo study the application of non-real-time ultrasound bronchoscopy combined with Metagenomic Next-Generation Sequencing (mNGS) for diagnosis in focal pulmonary infectious diseases. MethodsProspective inclusion of patients with focal pulmonary infection were randomly divided into two groups, the experimental group used non-real-time ultrasound bronchoscopy positioning to collect bronchial alveolar lavage fluid (BALF), while the control group used chest CT position. BALF was subjected to mNGS and traditional microbial detection including traditional culture, the fungal GM test and Xpert (MTB/RIF). ResultThe positive rate of traditional culture (39.58% vs. 16.67%, P=0.013) and mNGS (89.58% vs. 72.92%, P=0.036) in experimental group was higher. The positive rate of Xpert MTB/RIF (4.17% vs. 2.08%, P=1) and fungal GM test (6.25% vs. 4.17%, P=0.765) was similar. The positive rate of bacteria and fungi detected by mNGS was higher than traditional culture (61.46% vs. 28.13%, P<0.001). Mycobacterium tuberculosis was similar to Xpert MTB/RIF (8.33% vs. 3.13%, P=0.21). Aspergillus was similar to GM test (7.29% vs. 5.21%, P=0.77). The total positive rate of traditional microbial methods was 36.46%, but 81.25% in mNGS (P<0.001). mNGS showed that 35 cases were positive and 13 kinds of pathogens were detected in control group, but 43 patients and 17 kinds of pathogens were detected in experimental group. The average hospitalization time [(12.92±3.54) days vs. (16.35±7.49) days] and the cost [CNY (12209.17±3956.17) vs. CNY (19044.10±17350.85)] of experimental group was less (P<0.001). ConclusionsNon-real-time ultrasound bronchoscopy combined with mNGS can improve the diagnostic rate of focal pulmonary infectious diseases which is worthy of popularization and application in clinical practice.