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Table 1 Advantages and disadvantages of reported common methods for monitoring the aptamer-target binding in SELEX rounds

From: SELEX tool: a novel and convenient gel-based diffusion method for monitoring of aptamer-target binding

Monitoring methods

Suitable targets

Advantages

Limitations

References

Dot blotting

Protein

Focus on the candidate aptamers binding phase, which shows the real enrichment in SELEX;

Relative ease of performance

Sequence labeling required;

Not suitable for the target molecules with the same electrostatic charge as NC membrane used

[34]

qPCR

Protein & Small molecule

Relative ease of performance

Focus on the elution phase, which could be confused by non-specific eluates;

Error of nonspecific amplification;

[35,36,37,38]

EMSA

Protein

Focus on the candidate aptamers binding phase, which shows the real enrichment in SELEX

Sequence labeling required;

Labor- and time-consuming;

Not suitable for small molecule targets

[39]

Gel-shifting

Protein

Focus on the candidate aptamers binding phase, which shows the real enrichment in SELEX;

Easy for performance

Detection in none-binding conditions, I.E. electrophoretic buffer solution;

Not suitable for small molecule targets

[34, 36]

ELONA

Protein

Focus on the candidate aptamers binding phase, which shows the real enrichment in SELEX;

Relative ease of performance

Sequence labeling required;

Labor- and time-consuming;

Non-specific binding of candidates to the plate, which confuse the enrichment;

Not suitable for small molecule targets

[40]

Agarose gel analysis

Protein & Small molecule

Relative ease of performance

Focus on the elution phase, which could be confused by non-specific eluates;

Error of nonspecific amplification

[41]

HTS

Protein & Small molecule

Focus on the enrichment of candidate aptamers according to the SELEX rounds

Expensive cost required;

Focus on the elution phase, which could confuse by none-specific elution;

Time-consuming for sample preparation

[35, 42]

SPR

Protein & Small molecule

Focus on the candidate aptamers binding phase, which shows the real enrichment in SELEX

Expensive sensor chip required;

Labor- and time-consuming

[42, 43]

UV quantification

Small molecule

Ease of performance

Focus on the elution phase, which could confuse by none-specific elution

[44]

Fluorescence quantification

Small molecule

Relative ease of performance

Sequence labeled with fluorophore required;

Focus on the elution phase, which could confuse by none-specific elution

[45, 46]

Fluorescence binding assay

Small molecule

Focus on the candidate aptamers binding phase, which shows the real enrichment in SELEX;

Relative ease of performance

The autofluorescence of target required

[47, 48]

Gel-elution assay

Small molecule

Focus on the candidate aptamers binding phase, which shows the real enrichment in SELEX

Target-coupled column required;

Labor- and time-consuming

[49]

GBDM

Protein & Small molecule

Focus on the candidate-target binding phase, which shows the real enrichment in SELEX;

Easy for performance without expensive equipment;

Suitable for monitoring every selection step during SELEX process

Overnight diffusion required

Optimized in This work

  1. Notes: qPCR real-time quantitative PCR, EMSA Electrophoretic mobility shift assay, ELONA enzyme-linked oligonucleotide assay, HTS High throughput sequencing, SPR Surface plasmon resonance, GBDM gel-based diffusion method