Rhamnose

The polymeric copies of a rhamnose motif form glycopolymers (ARGPs), which can bind endogenous antirhamnose antibodies present in human serum, thus attract antibodies from human serum to the target cell surface.^[1]

Jurkat T cells are resuspended in the RPMI medium without FBS at a final concentration of 350000 cells/mL and placed on ice. After 30 min of incubation on ice, 1 mL of cold cell suspension is incubated for 1 h on ice in the dark with 40 μL of a 1 mg/mL stock solution of the CholA-p(RhamEAmAIBN) polymer in PBS. As a negative control, 1 mL of cold cell suspension is incubated for 1 h on ice in the dark with 40 μL of a 1 mg/mL stock solution of the CholA-p(GalEAmAIBN) polymer in PBS. After the incubation, cells are washed twice with PBS by centrifugation for 3 min at 500 G. Afterward, cells are resuspended in 500 μL of flow incubation buffer (1% bovine serum albumin, 5 mM Na2EDTA in PBS) containing 10% of human serum and are then incubated for 30 min in the dark on ice. After this incubation period, cells are centrifuged at 500 G for 3 min and washed once with PBS. Cells are spun down again at 500 G for 3 min and incubated during 1 h in the dark at room temperature with 500 μL of goat antihuman IgG secondary antibody bound to Alexa 647 at a final concentration of 4 μL/mL and 1% human albumin serum in PBS. Incubation is followed by two washing steps with PBS by centrifugation at 500 G for 5 min. Cells are imaged using stochastic optical reconstruction microscopy.

Rhamnose(Rha) conjugated immunogens induces high titers of anti-Rha antibodies in wildtype mice, which can replace α1,3galactosyltransferase knockout (α1,3GT KO) mice in such antigen/antibody-mediated vaccine design when developing cancer immunotherapies.^[2]