AptaSel - International training on SELEX (Initial + Advanced)

  • Curso abierto
  • Curso presencial

Ficha Técnica

Temario del Curso

Módulo:
Hands-on PCR
Duración:
7 h.
Contenido:

Hands-on PCR

    Theory (2 hours):
  • - Introduction to PCR (Polymerase Chain Reaction)
  • - Primer design and how to avoid primer dimer formation
  • - Design of a random DNA library
  • - PCR optimization of a random library
  • - Gel electrophoresis system and analysis of PCR data

    Practical (5 hours):
  • Hands-on lab running PCR and analysis of PCR by gel electrophoresis system

Módulo:
Target preparation
Duración:
7 h.
Contenido:

Target preparation

    Theory (2 hours):
  • - Targets, principal requirements for selection
  • - Characterization of selected target
  • - Partitioning strategies
  • - Immobilization of target, conjugation strategies
  • - Quality control

    Practical (5 hours):
  • Hands-on lab SDS-PAGE (Sodium Dodecyl Sulfate PolyAcrylamide Gel Electrophoresis), different immobilization and conjugation strategies, competitive ELISA (Enzyme Linked ImmunoSorbent Assay) -bead assay

Módulo:
Systematic Evolution of Ligands by EXponential Enrichment (SELEX)
Duración:
7 h.
Contenido:

    Theory (2 hours):
  • - Introduction to SELEX
  • - Different SELEX strategies
  • - Incubation and optimal conditions
  • - PCR, PCR optimization of a random library
  • - Preparation of single stranded DNA

    Practical (5 hours):
  • - Hands-on lab running SELEX (incubation, PCR, single stranded DNA preparation).

Módulo:
Evolution, Next Generation Sequencing (NGS), Truncation studies
Duración:
7 h.
Contenido:

This part covers the monitoring of evolution during SELEX, which allows control and adjustment of the selection pressure and stringency to achieve the desired properties of the selected aptamers. Next Generation Sequencing and data analysis with the interpretation of results and finally determination of secondary structure and truncation studies will be also discussed.

    Theory (2 hours):
  • - Monitoring of evolution during SELEX
  • - NGS and data analysis
  • - Truncation studies

    Practical (5 hours):
  • Hands-on lab evolution studies (APAA (Apta PCR affinity assay), SPR (Surface Plasmon Resonance), training on analysis of NGS data and truncation studies.

Módulo:
Binding assays or Lateral Flow Assays (LFAs)
Duración:
7 h.
Contenido:

Option A) Binding assays

The different binding assays that are routinely used to study the specificity and affinity of aptamers are described and the suitability for different types of binding assays for various types of targets is described.

    Theory (2 hours):
  • - Different binding assays: SPR, SPRi (Surface Plasmon Resonance imaging), APAA, ELAA (Enzyme Linked aptamer Assay), EMSA (Electrophoretic Mobility Shift Assay, MST (MicroScale thermophoresis) and BLI (BioLayer Interferometry).
  • - Suitability of binding assays to specific targets


    Practical (5 hours):
  • Hands-on lab binding assays (SPR, SPRi, APAA, ELAA, EMSA, MST and BLI)


Option B) Lateral Flow Assays (LFAs)

An introductory overview of lateral flow assays is provided, where the essential components of these assays and their importance is explained. Fundamental assay considerations and the implementation of aptamers in different lateral flow assay formats will be described.

    Theory (2 hours):
  • Introduction to aptamer based LFAs


    Practical (5 hours):
  • Hands-on lab competitive/sandwich aptamer based LFAs

Inscripción