Polymerase chain reaction (PCR) technique is widely used in many experimental conditions, and Taq DNA polymerase is critical in PCR process. In this article, the Taq DNA polymerase expression plasmid is reconstructed and the protein product is obtained by rapid purification, (“Rapid purification of high-activity Taq DNA polymerase” (Pluthero, 1993 [1]), “Single-step purification of a thermostable DNA polymerase expressed in Escherichia coli” (Desai and Pfaffle, 1995 [2])). Here we present the production data from protein expression and provide the analysis results of the production from two different vectors. Meanwhile, the purification data is also provided to show the purity of the protein product.

Lymph node metastasis is associated with poor prognosis of oral squamous cell carcinoma (OSCC), and few studies have explored the relevance of postoperative lymphatic drainage (PLD) in metastatic OSCC. Alpha-enolase (ENO1) is a metabolic enzyme, which is related to lymphatic metastasis of OSCC. However, the role of ENO1 in PLD in metastatic OSCC has not been elucidated. Herein, we collected lymphatic drainage after lymphadenectomy between metastatic and non-metastatic lymph nodes in OSCC patients to investigate the relationship between ENO1 expression and metastasis, and to identify the proteins which interacted with ENO1 in PLD of patients with metastatic OSCC by MS/GST pulldown assay. Results revealed that the metabolic protein apolipoprotein C-III (ApoC3) was a novel partner of ENO1. The ENO1 bound to ApoC3 in OSCC cells and elicited the production of interleukin (IL)-8, as demonstrated through a cytokine antibody assay. We also studied the function of IL-8 on Jurkat T cells co-cultured with OSCC cells in vitro. Western blot analysis was applied to quantitate STAT3 (signal transducer and activator of transcription 3) and p-STAT3 levels. Mechanistically, OSCC cells activated the STAT3 signaling pathway on Jurkat T cells through IL-8 secretion, promoted apoptosis, and inhibited the proliferation of Jurkat T cells. Collectively, these findings illuminate the molecular mechanisms underlying the function of ENO1 in metastasis OSCC and provide new strategies for targeting ENO1 for OSCC treatment.

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Background. Adoptive cell therapy (ACT) based on chimeric antigen receptors (CARs) expressed on the surface of T cells shows a remarkable clinical outcome, particularly for B-cell malignancies. However, toxicity and side effects of CD19-redirected CAR T cells have been observed concurrently in most cases due to cytokine release and tumor cell lysis. Therefore, strictly controlling the amount of valid T cells re-transfused to patients seems to be an important step in reducing toxicity and side effects of CAR T cells. Transfection efficiency via lentiviral particles varies widely in different cases.Objectives. The aim of this study was to accurately calculate and control the number of valid CAR T cells through ACT because the restriction antibiotics gene or the fluorescence gene are not suitable for tracking or screening for valid transfected T cells. Material and methods.We expressed and purified a GFP-CD19 fusion protein as a probe to measure the expression efficiency of CD19-redirected CAR on the cell surface in adherent and suspension cell lines. Results.We can precisely calculate the transfected efficiency of lentiviral particles by counting the number of GFP-labeled cells under a microscope, as well as calculate the percentage by comparing the number of GFP-labeled cells to total cells. Conclusions.We propose a method to control the number of valid cells in ACT and to reduce toxicity and side effects in clinical use -a convenient technique for monitoring the dosage of CAR T cells for patients.