Acknowledgements:

Thank

• My sponsor
• My family
• Professor Denise Jackson, Laboratory Medicine Program Leader
• Fiona Morris, Project Demonstrator

Method 1.

Monolayer formation and stability:

1. A) Monolayer Formation:

• Make 0.2% red cell suspension from each panel cell (3% red cell suspension) by adding 40 µL of panel cell to 560 μL diluent (CFB)
• Add 50 µL of 0.2% red cell suspension to the corresponding well in the micro titer plate
• Centrifuge the plate at 350 g for 3 minutes

Ref: Llopis F, Carbonell‐Uberos F, Planelles M, Montero M, Plasencia I, Carrillo C. A new microplate red blood cell monolayer technique for screening and identifying red blood cell antibodies. Vox sanguinis. 1996;70(3):152-6.

1. B) Monolayer Stability (Antibody Detection by Solid phase red cell assay):

Sample A. serum with known Anti-D (IgG antibody)

• Add 100 µL of LISS to each well
• Add 100 µL of test serum to each well
• Incubate the plate at 37 ᵒC for 20 minutes
• During the incubation prepare 50% (½ strength) AHG reagent (diluted in NBS) and 0.2% AHG control
• After the incubation wash the plate 5 times with NBS
• Add 50 µL of 50% (½ strength) AHG reagent
• Add 50 µL of 0.2% AHG control cells
• Centrifuge the plate at 500 g for 2 minutes and read

Ref: Llopis F, Carbonell‐Uberos F, Planelles M, Montero M, Plasencia I, Carrillo C. A new microplate red blood cell monolayer technique for screening and identifying red blood cell antibodies. Vox sanguinis. 1996;70(3):152-6.

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