TITLE: ChroPlate - ProteinA
AUTHORS: Alexandra Ehl, David Frommholz, Nadine Stefanczyk

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span style = "font-weight:bold;">Purification Guide for the Isolation of Antibodies with ChroPlate Filtration Plates by DALEX Biotech.</span></div><div class = "text-block">Easy and quick high throughput antibody purification from various sources and species.</div><div class = "text-block">Each well of the ChroPlate has a binding capacity of > 1 mg (tested with human polyclonal Ig, binding varies between species and clones).</div><div class = "text-block">The proprietary resin does not shrink or swell in aqueous buffers.</div><div class = "text-block">High pressure stability.</div><div class = "text-block">pH stability short term 2 - 8, long term 3 - 8.</div><div class = "text-block">Excellent thermal stability up to 15 minutes at 80 °C in aqueous buffers at neutral pH.</div><div class = "text-block">Can be dried for long term storage (80 °C for > 2 h).</div></div>

[STEPS]
?. [Load and Wash]
Empty the deep-well plate or place the ChroPlate on a clean one. Add 500 µl wash buffer to each well and centrifuge 2 minutes at 400 g in a swing-out rotor.
In case of unspecific hydrophobic and/or ionic interactions include up to 1 % Tween-20 and/or up to 0.5 M NaCl in the wash buffer.
?. [Elution]
Place the ChroPlate on a clean deep-well plate. Add 300 µl elution buffer to each well and centrifuge 2 minutes at 400 g in a swing-out rotor.Repeat two more times.
?. [Equilibration]
Add 500 µl wash buffer to each well, place the ChroPlate on top of a deep-well plate, and centrifuge 2 minutes at 400 g in a swing-out rotor.  For counterbalance of the centrifuge a dummy filter plate is included in the kit.
?. [Load and Wash]
Repeat the previous step.
For increased purity, repeat the washing step a third time.
?. [Elution]
Add one drop of neutralization solution to every well of the deep-well plate and mix gently.
?. [How do you want to purify?]
It is advisable to purify by centrifugation and not by vacuum filtration because of excessive frothing.
?. [Equilibration]
Assemble your vacuum device according to the manufacturer's instructions.Add 500 µl wash buffer to each well, place the ChroPlate on a waste receptacle and apply a vacuum of 100 mbar for 2 minutes.
?. [Load and Wash]
Place the ChroPlate on a clean deep-well plate. Add up to 1.5 ml sample to every well. Apply a vacuum of 100 mbar for 2 - 3 minutes.
It is advisable to keep the flow-through and wash fractions for later analysis, e.g. SDS-PAGE.Strength and time of the vacuum depends on the sample's volume and viscosity. Volumes larger than 1 ml and viscous samples e.g. serum usually require more than 2 minutes.For optimal binding and purity, the pH of the sample should be 7.5 - 8.5 and should contain 150 - 300 mM NaCl. An easy way to achieve this is by adding 1/11 volume 0.5 M Tris, 2 M NaCl (pH 8.0) to your sample.
?. [Load and Wash]
Empty the deep-well plate or place the ChroPlate on a clean one. Add 500 µl wash buffer to each well and apply a vacuum of 100 mbar for 2 minutes.
In case of unspecific hydrophobic and/or ionic interactions include up to 1 % Tween-20 and/or up to 0.5 M NaCl in the wash buffer. However, frothing appears easily when using Tween in vacuum filtration procedures.
?. [Load and Wash]
Repeat the previous step.
For increased purity, repeat the washing step a third time.
?. [Elution]
Place the ChroPlate on a clean deep-well plate. Add 100 µl elution buffer to each well apply a vacuum of 100 mbar for 2 minutes.Repeat two more times.
?. [Elution]
Add one drop of neutralization solution to every well of the deep-well plate and mix gently.
?. [Load and Wash]
Place the ChroPlate on a clean deep-well plate. Add up to 1.5 ml sample to every well. Centrifuge 5 minutes at 400 g in a swing-out rotor.
It is advisable to keep the flow-through and wash fractions for later analysis, e.g. SDS-PAGE.The centrifugation time depends on the sample's volume and viscosity. Volumes larger than 1 ml and viscous samples e.g. serum usually require more than 5 minutes centrifugation time.For optimal binding and purity, the pH of the sample should be 7.5 - 8.5 and should contain 150 - 300 mM NaCl. An easy way to achieve this is by adding 1/11 volume 0.5 M Tris, 2 M NaCl (pH 8.0) to your sample.