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

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span style = "font-weight:bold;">Purification Guide for the Isolation of Antibodies with ChroDrip Columns by DALEX Biotech.</span></div><div class = "text-block">Easy and quick small scale antibody purification from various sources and species.</div><div class = "text-block">Each ChroDrip column has a binding capacity of > 10 mg/ml (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]
Add your sample to the top of the column and let it flow through by gravity.
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.Removal of particulate matter from the sample by centrifugation or filtration (0.45 µm) is recommended.For slow or weak binding antibodies collect the flow through and apply it again.
?. [Load and Wash]
Add 5 column volumes of wash buffer and wait until it has drained. Repeat once more.
It is advisable to keep the flow-through and wash fractions for later analysis, e.g. SDS-PAGE.For increased purity, repeat the washing step up to 5 times.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]
Add 0.75 column volumes of elution buffer to the column.
This fraction does not contain the target protein. The small amount of elution buffer replaces most of the wash buffer in the column. This "pre-elution step" will result in a more concentrated eluate.
?. [Elution]
Place a clean tube under the column. Add 3 times 3 column volumes of elution buffer to the column, collect each fraction in a separate tube. Wait inbetween the elution steps until the buffer has drained completely.
For more concentrated eluates, elute 8 times with one column volume and collect each fraction into a separate tube. Determine in which fractions most of the protein is contained and combine these.
?. [Equilibration]
Remove the bottom and top cap of the column and add 5 column volumes wash buffer (bed volume is written on the column) to the column.
If you work with a used column, drain the storage solution first.
?. [Cleaning and Storage]
Wash the column successively with 5 column volumes elution buffer, 5 column volumes wash buffer and 5 column volumes deionized water. Then, add 10 column volumes 20 % ethanol or wash buffer (contains 0.05 % (w/v) sodium azide). Wait until half of the buffer has drained. Close the top lid and then the bottom stopper. Store at room temperature or at 4 - 8 °C.Alternative for long-term storage:Dry the open (top and bottom) column in an oven at 80 °C for at least 2 hours or over night. Make sure the bottom stopper is completely dry, too. Put on the column's outlet, close the lid and store the column closed at room temperature.
?. [Elution]
Add neutralization solution to the eluate. For each milliliter of eluate add three drops of neutralization solution.
?. [What do you want to do?]
Do you want to purify antibodies or sanitize your column?Please choose below.
?. [Sanitization]
After a purification: wash the column with 5 column volumes of elution buffer and 5 column volumes of wash buffer. Let the buffer drain.If the column was stored: let the buffer drain and wash with 5 column volumes of wash buffer.
?. [Sanitization]
Add 5 column volumes of sanitization solution, let 1 column volume drain and then close the column with the top and bottom cap. Incubate for one hour at RT.
?. [Sanitization]
Drain the sanitization solution and wash the column with 10 column volumes of deionized water.
?. [Storage]
Add approximately 5 column volumes of wash buffer (contains 0.05 % (w/v) sodium azide). Wait until half of the buffer has drained. Close the top lid and then the bottom stopper. Store at room temperature or at 4 - 8 °C.It is recommended to wait at least 12 hours before the next purification.Alternative for long-term storage:Dry the open (top and bottom) column in an oven at 80 °C for at least 2 hours or over night. Make sure the bottom stopper is completely dry, too. Put on the column's outlet, close the lid and store the column closed at room temperature.