Nanoantibodies (single-domain antibodies, nanobodies) derived from noncanonical single-chain immunoglobulins provide an attractive tool for in vitro and in vivodiagnostics as well as for development...

Alternative splicing is one of the main regulation pathways in living cells beyond simple changes in the level of protein expression. Most of the approaches proposed in proteomics for the identification of specific splicing isoforms require a preliminary deep transcriptomic analysis of the sample under study, which is not always available, especially in the case of the re-analysis of previously acquired data. Herein, we developed new algorithms for the identification and validation of protein splice isoforms in proteomic data in the absence of RNA sequencing of the samples under study. The bioinformatic approaches were tested on the results of proteome analysis of human melanoma cell lines, obtained earlier by high-resolution liquid chromatography and mass spectrometry (LC-MS). A search for alternative splicing events for each of the cell lines studied was performed against the database generated from all known transcripts (RefSeq) and the one composed of peptide sequences, which included all biologically possible combinations of exons. The identifications were filtered using the prediction of both retention times and relative intensities of fragment ions in the corresponding mass spectra. The fragmentation mass spectra corresponding to the discovered alternative splicing events were additionally examined for artifacts. Selected splicing events were further validated at the mRNA level by quantitative PCR.

This article describes a method for activating lymphocytes isolated from the peripheral blood of melanoma patients and cultured in a medium supplemented with IL‑2 and IL‑15. It was shown that in these terms, lymphocytes have an increased proliferative and activation potential. The combination of cytokines has a positive effect on cytotoxicity, viability and the expression of activation markers (CD38, CD69, CD25, HLA-DR and NKG2D) on NK- and T-lymphocyte, and may be recommended for the culture of lymphocytes in melanoma patients for the purpose of adoptive immunotherapy.