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兔单抗亲和力和灵敏度更高
相对于鼠单抗（解离常数Kd在10-9-10-10M水平），兔单抗的亲和力可提高100-1000倍（Kd在10-10-10-12M水平）。实际使用时，兔单抗的工作浓度更低，灵敏度更高，因此产生的背景更低，大大降低了假阳性的发生。用作免疫组化等用途时，有时甚至可免去抗原修复的步骤。
兔单抗能识别更多表位
很多蛋白在人类和啮齿动物中同源性很高，这些保守表位在小鼠体内免疫原性很弱，不容易产生高质量抗体。而兔做为宿主，可以更好地识别这些表位，产生针对更多表位的抗体。当然，兔也适合生产针对鼠类蛋白的抗体。
Fusion and Cloning
Reagents
(StemCell Technologies, Inc. # 03800)
Medium A - Pre-fusion Medium and Hybridoma Expansion Medium (StemCell Technologies, Inc. - # 03801)
Medium B - Fusion Medium (StemCell Technologies, Inc. - # 03802)
Medium C - Hybridoma Recovery Medium (StemCell Technologies, Inc. - # 03803)
Medium D - Hybridoma Selection Medium (StemCell Technologies, Inc. - # 03804)
Medium E - Hybridoma Growth Medium (StemCell Technologies, Inc. - # 03805)
PEG Solution (StemCell Technologies, Inc. - # 03806)
Materials
50 ml Sterile conical tubes (Falcon #2070)
15 ml Sterile conical tubes (Falcon #2099)
10 ml sterile pipets (Falcon # 7551)
1 ml sterile pipets (Falcon #7521)
Pasteur Pipets, sterile
100 mm sterile Petri Dishes (Falcon #1009)
96-well culture dishes (Falcon # 3072)
24-well culture dishes (Falcon # 3047)
Forceps
Scissors
Multi-channel pipettor, 50-200 ml
Pipet tips, sterile
Reagent Reservoir, sterile
Tupperware container
Myeloma Cells
One week prior to the fusion, split myeloma cells into Medium A to ensure that they are well adapted.
Grow up approximately 2 x 107 healthy cells, in mid-log phase, for each fusion.
Fusion
Count the myeloma cells and resuspend to 2 x 107 cells in 30 ml Medium A in a 50 ml tube.
Sacrificed the mouse, saturate in ethanol, and remove the spleen.
Place the spleen in a Petri dish containing 10 ml of Medium A.
Prepare a single cell suspension of the spleen.
Using a Pasteur Pipet, transfer the spleen cells to a 50 ml tube.
Rinse the Petri dish with another 10 ml of Medium A and add to the tube.
Allow the tube to sit for approximately 1 minute to settle the larger pieces of tissue. Transfer the cell suspension to a clean tube, leaving behind the larger pieces of tissue.
Add 10 ml of Medium A to the tube to wash the tissue pieces. Allow to settle. Transfer the medium to the clean tube, combining it with the previous cell suspension.
Centrifuge the splenocyte suspension at 400 g for 10 minutes, RT.
Resuspend the cells in 10 ml of Medium A and count.
Combine 108 viable spleen cells with 2 x 107 myeloma cells in a 50 ml tube. Centrifuge at 400g for 10 minutes.
Discard the supernatant and wash the pellet twice with 40 ml Medium B, pre-warmed to 37oC.
Discard the supernatant. Tap the bottom of the tube to loosen the pellet.
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