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Preparation method and application of pMHC polymer based on DNA nanotechnology

A nanotechnology and institutional technology, applied in the field of T cell detection, can solve the problems of insufficient binding affinity and unsatisfactory performance, and achieve the effect of cost saving and high efficiency

Pending Publication Date: 2022-06-21
THE NAT CENT FOR NANOSCI & TECH NCNST OF CHINA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, pMHC tetramer technology still has some limitations
Due to insufficient binding affinity, pMHC tetramers in antigen-specific CD4 + T cells, low-affinity αβ T cells, and rare γδ T cells perform poorly in assays

Method used

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  • Preparation method and application of pMHC polymer based on DNA nanotechnology
  • Preparation method and application of pMHC polymer based on DNA nanotechnology
  • Preparation method and application of pMHC polymer based on DNA nanotechnology

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] Example 1 PMHC multi -gathering system preparation method based on DNA nanotechnology

[0055] This example provides a precise space assembly method and maximum assembly efficiency test based on DNA nano -technology control PMHC monomer.

[0056] First, SPYTAG, which is modified by clicked by clicking a chemical reaction to connect with the complementary chain containing DBCO, and use 100kda centric filters to purify the product.

[0057] Secondly, the SPYTAG-DNA complementary chain and PE modified complementary chain and rectangular DNA nano structure (with the corresponding capture chain) are annealing in the PCR instrument.And purified through 100kda centric filters to remove excess proteins.Among them, the preparation method with a rectangular DNA nanogenic structure with a specific capture chain is as follows: 1 template chain and several short chains are paired with a complementary pairing of alkali base to form a rectangular DNA nano structure with a specific capture...

Embodiment 2

[0070] Example 2 peptide based on DNA nanotechnology-Main tissue compatibility complex (DNA-PMHC) polyette detection and analysis of specific T cell efficiency assessment

[0071] In order to test the detection efficiency of the specific T cells of the present invention, C57BL / 6 mice spleen cells were selected for testing.

[0072] First, use OVA 257-264 (SIINFEKL) antigen peptides and Polyicica (IC) mixture to 1: 1 subcutaneous immune C57BL / 6 mice, immune every 3 days for immune 3 times.After 7 days, remove the spleen and prepare the spleen single -cell suspension.

[0073] Second, use H-2K b -OVA 257-264 (SIINFEKL) Teti-cluster as a control and DNA nanogenic polymerizer for dyeing H-2K, respectively b Limited OVA257-264 specific TCR.

[0074] Finally, use the current cytometer 2060R, Aglen) to measure the sample, and use Novoexpress to analyze data.

[0075] The experimental results show that the use of DNA-PMHC to detect antigen-specific T cells. The number of PMHCs from 3-12 D...

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Abstract

The invention discloses a preparation method and application of a pMHC polymer based on a DNA nanotechnology. According to the invention, the DNA nanostructure is used as a support to connect different numbers of pMHCs, the number, position and direction of the pMHCs are controlled, and an atomic force microscope is used to clearly observe that the different numbers of pMHCs are captured and displayed on the DNA nanostructure. The DNA-pMHC is used for detecting the antigen-specific T cells, and due to the unidirectional multivalence effect mediated by the single orientation of the pMHC on the DNA nanostructure, the sensitivity of a DNA-pMHC multimer with the number of pMHC ranging from 3 to 12 is 1.9-4.3 times that of a pMHC tetramer when the antigen-specific T cells are detected is 1.9-4.3 times that of the DNA-pMHC multimer with the number of pMHC ranging from 3 to 12. The pMHC polymer based on the DNA nanotechnology has wide application prospects in analysis of antigen-specific T cells, and a powerful tool is provided for exploring the spatial biological effect of accurate protein arrangement under the nanoscale.

Description

Technical field [0001] The present invention belongs to the field of T cell testing. Specifically, it involves a PMHC multi -gathering system and application method based on DNA nanotechnology. Background technique [0002] CTL is the main effect cells of the body's mediated cellular immunity and the core of adaptive cell immunity.It plays an important biological role in reactions such as the body's intracellular pathogen infection, anti -tumor and rejection of transplantation.The antigen specialties CTL is a small group of important cells, but because they are very low in lymphocytes, it is very difficult to detect this group of important cells.To accurately monitor the progress of the disease and evaluate the effectiveness of the vaccine requires efficiency, specificity and quantitative detection of antigen -specific T cells. [0003] Synthetic peptide-major tissue composite complex (PMHC) can be combined with T cell receptor (TCR) on the surface of antigen-specific T cells. Th...

Claims

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Application Information

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IPC IPC(8): C07K14/74C07K1/107G01N33/569G01N33/543G01N33/533
CPCC07K14/70539G01N33/56977G01N33/54346G01N33/533G01N2333/70539
Inventor 赵潇马娜娜
Owner THE NAT CENT FOR NANOSCI & TECH NCNST OF CHINA
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