[Cloning, expression and purification of arginine kinase from Blattella germanica and its immune activity].

OBJECTIVE: To clone and express the arginine kinase (AK) gene of Blattella germanica and analyze its immune activity. METHODS: The cDNA of AK was cloned using specific primers from the total RNA of Blattella germanica The open reading frame (ORF) of AK was cloned into pET-28A vector, and expressed in Escherichia coli BL21(DE3) with IPTG induction. The recombinant protein was purified by Ni2+ chelating affinity chromatography. The recombinant protein was detected by SDS-PAGE, and its immune activity was analyzed by Western blotting. RESULTS: The cloned cDNA ORF sequence (GenBank accession No. FJ514482) contained 1071bp and encoded 356 amino acids. Its sequence homology with the published one (GenBank accession No. EU429466) was 97.2% at nucleotide level. The recombinant containing recombinant plasmid pET-28a-AK expressed a soluble protein of AK (Mr 45 000) after being induced with IPTG. The recombinant AK protein was recognized by sera of allergic patients, indicating that the recombinant AK protein has an adequate response activity. CONCLUSION: The AK gene of Blattella germanica has been cloned and the recombinant AK protein has been confirmed with immune activity.

Lymphocyte subsets immune characteristics in hand foot and mouth disease：Its detection and clinical value / 上海预防医学

Objective] To summarize the immune characteristics in hand foot and mouth disease (HFMD) children with regards to peripheral blood T lymphocyte , B lymphocyte, NK cell and NKT cell, and to discuss its mechanism in the development of HFMD and their detection and clinical value . [ Methods] By means of flow cytometry , 152 children with HFMD and 45 healthy children were detected for the percentage of T lymphocyte , B lymphocyte, NK cell, NKT cell in peripheral blood . [ Results] The percentage of CD3 +,CD3 +CD4 +, CD3 +CD8 +, NK cells and ratio of CD4 +/CD8 +of HFMD acute phase group were lower than those in healthy group , while NKT cells, B cells were higher.The percentage of CD3 +, CD3 +CD4 +, CD3 +CD8 +, NKT cells and CD4 +/CD8 +ratio of HFMD acute phase were lower than those of recovery phase , while B cells and NK cells were higher .The percentage of CD 3 +cells, CD3 +CD4 +cells, CD3 +CD8 +cells, NKT cells and the ratio of CD 4 +/CD8 +in severe HFMD group were lower than those in mild group , while B cells and NK cells were higher . [ Conclusion] The immune function of children with HFMD proved disordered;CD3 +cells, CD3 +CD4 +cells and B cells in peripheral lympho-cytes were increased during the acute phase of HFMD;NKT cells increased during recovery phase of HFMD.We conclude that lymphocyte subsets detection can be used as an index of prognosis of HFMD in children .