Fc-specific and covalent conjugation of a fluorescent protein to a native antibody through a photoconjugation strategy for fabrication of a novel photostable fluorescent antibody.

Fluorophore-antibody conjugates with high photobleaching resistance, high chemical stability, and Fc-specific attachment is a great advantage for immunofluorescence imaging. Here, an Fc-binding protein (Z-domain) carrying a photo-cross-linker (p-benzoylphenylalanine, Bpa) fused with enhanced green fluorescent protein (EGFP), namely photoactivatable Z-EGFP recombinant, was directly generated using the aminoacyl-tRNA synthetase/suppressor tRNA technique without any further modification. By employing the photoactivatable Z-EGFP, an optimal approach was successfully developed which enabled EGFP to site-selectively and covalently attach to native antibody (IgG) with approximately 90% conjugation efficiency.

Chest Computed Tomography Image for Accurately Predicting the Optimal Insertion Depth of Left-Sided Double-Lumen Tube.

Objective: The main objective of this study was to assess the feasibility and accuracy of measuring the distance between the vocal cord and carina using chest computer tomography (CT) as a guide for the intubation of a left-sided double-lumen tube (LDLT).

Design: Single-center, prospective, randomized study.

Setting: Local hospital in China.

A comparison of extraluminal and intraluminal use of the Uniblocker in left thoracic surgery: A CONSORT-compliant article.

Background: The aim of this study was to assess the feasibility and safety issues concerning extraluminal use of the Uniblocker for one-lung ventilation (OLV) in the left thoracic surgery.

Methods: Forty patients undergoing elective left thoracic surgery were included in this study, and all patients were randomly allocated to extraluminal use of Uniblocker group (E group, n = 20) or intraluminal use of Uniblocker group (I group, n = 20). Time for intubation, time for verification of the correct position of Uniblocker, incidence of Uniblocker displacement, index of pulmonary collapse, mean arterial pressure, heart rate, peak airway pressure, oxygen saturation in two-lung ventilation, and 30 minutes after OLV, bronchial damage after OLV, sore throat, and hoarseness postoperative were recorded.

Immobilization of unraveled immunoglobulin G using well-oriented ZZ-His protein on functionalized microtiter plate for sensitive immunoassay.

Highly efficient protein immobilization is extremely crucial for solid-phase immunoassays. We present a strategy for oriented immobilization of functionally intact immunoglobulin G (IgG) on a polystyrene microtiter plate via iminodiacetic acid (IDA)-Ni(2+) and ZZ-His protein interaction. We immobilized a ZZ-EAP (Escherichia coli alkaline phosphatase)-His fusion protein, which exhibits Fc binding, His tag, and intrinsic AP activities, and analyzed it against the interaction between rabbit IgG anti-horseradish peroxidase (anti-HRP) and its binding partner HRP to investigate the specificity and efficacy of this method.

Comparative characterization of recombinant ZZ protein-alkaline phosphatase and its application in enzyme immunoassays.

A functional fusion protein, which consists of an antibody and an enzyme that can be used in enzyme immunoassays, has been constructed. However, a quantitative comparison of the characteristics of fusion proteins and chemical conjugates of the parents, which are functionally produced in a uniform microbial system, has not been adequately achieved. In this study, a fusion protein between the ZZ protein and Escherichia coli alkaline phosphatase (AP) and the parental ZZ protein and AP for chemical conjugate was functionally produced in the same bacterial system.

[The expression vector of murine secreting IL-1β promotes proliferation and migration of Hepa1-6 hepatoma cells].

Aim: To establish a murine secreted mature peptide IL-1β expression vector, transfect into Hepa1-6 hepatoma cells, and analyze the effect of recombinant IL-1β on proliferation, migration, and its specific expression in Hepa1-6 hepatoma cells.

Methods: The murine AFP signal peptide encoding sequence and mature IL-1β encoding fragment were linked together through overlapping PCR, and the chimeric DNA sequence was then inserted into a liver specific expression vector pLIVE(TM); to make a recombinant pLIVE-smIL-1β which expressed secreted murine IL-1β of classical pathway. pLIVE-smIL-1β, pLIVE(TM); and pLIVE-lacZ were transfected into Hepa1-6 by jetPEI respectively.

[IL-1β can promote proliferation and migration of Hepa1-6 cells and impair IFNα-induced cell growth inhibition in vitro].

Aim: To analyze the effect of recombinant IL-1β on proliferation, migration, and the effect on IFNα induced cell growth inhibtion.

Methods: The vector pLIVE-mIL-1β was transfected into Hepa1-6 cells mediated by transIT-LT1. Gene expression level of IL-1β was analyzed by RT-PCR and Sandwich ELISA.

[Preparation and characterization of monoclonal antibodies for human BCSC-1 protein].

Aim: To obtain monoclonal antibodies against human BCSC-1 protein for further study of the structure and function of human BCSC-1 protein.

Methods: pET-30a-BCSC-1 plasmid was constructed and transformed into E.coli BL21 (DE3) to express recombinant protein.

[Establishment of AFP promoter operated murine IL-1beta recombinant vector and its expression in H22 cells].

Aim: To establish a hepatoma specific murine IL-1beta (mIL-1beta) expression vector operated by AFP promoter and analysis its expression in H22 cell.

Methods: The chimeric operating sequence composed of the minimal AFP promoter and CMV enhancer(ECMV) was prepared through SOE-PCR. The sequence was inserted to replace the conventional enhancer and promoter in pIRES2-EGFP to establish the novel hepatoma specific vector p(afp)IRES2-EGFP.

[Expression of recombinant mouse IL-1beta significantly down-regulates NK cell mediated cytolysis [corrected] against H22 hepatoma cells].

Aim: To investigate the effect of the expression of recombinant IL-1beta in H22 hepatoma cells on its response to NK cell mediated cytotoxicity.

Methods: BALB/c mouse was stimulated by 6% of starch. Total RNA was prepared from peripheral blood monocytes (PBMCs).

[IL-1beta antisense RNA enhanced sensitivity of HepG2 cells to NK cell mediated cytotoxicity].

Aim: To investigate the inhibitory effect of IL-1beta antisense RNA on the sensitivity of HepG2 cells to the NK cell mediated cytotoxicity.

Methods: Two gene segments of IL-1beta [IL-1beta1(17-331, 315 bp) and IL-1beta2(246-505, 260 bp)] were selected for antisense RNA. Total RNA was extracted from PBMC of a healthy donor treated with LPS.