Immuno-affinity partition of cells in aqueous polymer two-phase systems

Area of Science:

• Bioconjugation Chemistry
• Immunology
• Biophysical Chemistry

Background:

• Antibodies are crucial for cell targeting but can cause non-specific aggregation.
• Poly(ethylene glycol) (PEG) modification is a common strategy to improve protein properties.
• Aqueous two-phase systems (ATPS) offer potential for cell separation but often lack resolution.

Purpose of the Study:

• To investigate the effect of poly(ethylene glycol) (PEG) modification on IgG antibodies targeting red blood cells.
• To assess the utility of PEG-modified antibodies in enhancing cell partitioning within dextran-PEG aqueous two-phase systems.
• To evaluate the efficiency of PEG-antibody modified cells in separation applications.

Main Methods:

• Covalent coupling of poly(ethylene glycol) (PEG) to anti-red blood cell IgG antibodies.
• Characterization of PEG-antibody conjugates, including molecular weight and degree of PEGylation.
• Utilizing dextran-PEG aqueous two-phase systems for cell partitioning and countercurrent extraction.
• Separation of mixed human and sheep red blood cells using PEG-derivatized antibodies.

Main Results:

• PEG modification reduced antibody-induced red blood cell agglutination.
• PEGylated antibodies exhibited increased affinity for the PEG-rich phase in ATPS.
• Red blood cells exposed to PEG-modified antibodies showed significantly enhanced partitioning into the PEG-rich phase.
• Complete separation of mixed red blood cells was achieved within 100 minutes using a 30-transfer countercurrent extraction.

Conclusions:

• Poly(ethylene glycol) modification is an effective strategy to improve antibody-based cell targeting and separation in aqueous two-phase systems.
• The degree of PEGylation and PEG molecular weight influence antibody properties and cell partitioning.
• This approach offers a highly efficient and specific method for separating red blood cells.