How to choose the TCR-Seq method? An in-depth evaluation of a 54-point SCI article is worthy of attention!

Perhaps you have also noticed that in recent years, more and more scientific researchers at home and abroad have devoted themselves to the research camp of immune repertoire

Among many TCR research methods, NGS technology has gradually become the mainstream analysis method of TCR

Related background of TCR-Seq

▪ Starting material DNA or RNA?

Most TCR-Seq methods can be roughly divided into DNA-based methods and RNA-based methods based on the starting material

▪ Database construction strategy

Two commonly used methods are mPCR (multiplex PCR) and 5'RACE (Rapid Amplification of cDNA Ends)

mPCR supports DNA and RNA initiation and usually includes two rounds of PCR

Literature interpretation

Four key factors need to be considered when choosing a TCR-Seq method: reproducibility, repeatability, sensitivity and specificity

Analyze the repeatability and reproducibility of different TCR-Seq methods

The research team compared the repeatability (ie, the similarity between data obtained by different samples using the same method) and reproducibility (ie, the similarity between data obtained by different methods) of different TCR-Seq

Analyze the sensitivity and specificity of different TCR-Seq methods

The next evaluation is for sensitivity and specificity

The dataset generated by RACE-3 contains up to 50% of TRA and up to 40% of TRB MRCs, compared with 10-20% of other RACE methods (Figure 5, Panel A in the literature)

in conclusion

In these TCR-Seq methods, the diversity and detection of rare clones are highly dependent on the type and quantity of starting materials

Editor's summary:

Although the Eugster team’s method was considered to be the most reproducible method for TRA analysis in the final analysis, the sensitivity of the SMARTer Human TCR a/b Profiling Kit is comparable to the Eugster team’s method and company A’s platform

Takara Bio has always been committed to providing our customers with high-quality and innovative products

SMARTer Human TCR a/b Profiling Kit v2 (TCRv2)

• Support 10 ng-1 μg total RNA derived from peripheral blood lymphocytes or 1 ng-100 ng T cells, as well as complete 1,000-10,000 T cells after purification;

I hope that the content shared in this issue can help you choose a suitable TCR-Seq library construction plan!

references

1.

Extended reading: 50 20-point articles, netizens: Immune repertoire analysis should be popular? !

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