Purification of Self-Assembling Protein Nanoparticles using Affinity Chromatography

Published: May 31, 2024

Abstract

Source: Karch, C. P. et al., Production of E. coli-expressed Self-Assembling Protein Nanoparticles for Vaccines Requiring Trimeric Epitope Presentation. J. Vis. Exp. (2019)

This video showcases the purification of histidine-tagged self-assembled protein nanoparticles or SAPNs using affinity liquid chromatography. The resulting purified SAPN fractions are suitable for vaccine development, holding promise for potential immunotherapeutic applications.

Protocol

1. Expression of the Self-assembling protein nanoparticles core containing a six-helix bundle (SHB-SAPN) Protein in E. coli BL21(DE3) Mix 95 mL of component A and 5 mL of component B of the media in a 2 L sterile glass Erlenmeyer flask as per the manufacturer's instructions (see the Table of Materials). Add ampicillin to a final concentration of 100 μg/mL. Inoculate the media with E. coli from a previously established glycerol stock cultur…

Açıklamalar

The authors have nothing to disclose.

Materials

2-propanol Fisher BP26181 4 L
Ampicillin Fisher BP1760-25 25 g
Corning Disposable Vacuum Filter/Storage Systems Fisher 09-761-108 A variety of sizes
GE Healthcare 5 mL HisTrap HP Prepacked Columns GE HealthCare 45-000-325 5 pack
Glycerol Fisher BP229-4 4 L
Imidazole Fisher O3196-500 500 g
Magic Media E. coli Expression Medium ThermoFisher K6803 1 L
MilliporeSigma Millex Sterile Syringe 0.22 mm Filters Millipore SLGV033RB 250 pack
One Shot BL21 Star (DE3) Chemically Competent E. coli ThermoFisher C601003 20 vials
Sodium Phosphate Monobasic Fisher BP329-500 500 g
Tris Base Fisher BP152-1 1 kg
Urea Fisher BP169-500 2.5 kg
Equipment
Fiberlite F14-14 x 50cy Fixed-Angle Rotor ThermoFisher 096-145075 Rotor
NGC Quest 10 Chromatography System BioRad 7880001 FPLC to aid in protein purification
Sonifer 450 Branson also known as 096-145075 Sonicator

Etiketler

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Bu Makaleden Alıntı Yapın
Purification of Self-Assembling Protein Nanoparticles using Affinity Chromatography. J. Vis. Exp. (Pending Publication), e22267, doi: (2024).

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