Recyclable CIM disks and papers

Another paper on anti-glycophorin-A MAb "purification" (compare to the one described in the post "Acid Test for antibody function?" by Ambur P. Dhivya's 2010 paper in Chromatographia)

CIM-EDA has been used in both papers.  In this latest one (2012), ~ 8 fold purification was seen with elution with 25 mM MOPS, pH 6.5 and 0.2 M NaCl.
In the older one (2010), 20 mM Tris buffer, pH 8, 0.6 M NaCl gave ~74 fold purification.

Images are still difficult to make out; but why change the conditions and report worse "purification" with different conditions?

abstract from Ambur Dhivya et al. 2010 Chromatographia

Dead cells producing antibody?

Side-by-side comparisons of chromatographic purification gels and profiles from the older (blue arrows and underlined in blue) Chromatographia 2010 paper and the recent (red arrows and underlined in red) Hybridoma 2012 paper.

Kumar BP, Rajak P, Vijayalakshmi MA, & Jayaprakash NS (2012). Production of Human Anti-Glycophorin-A Monoclonal Antibodies and Their Purification by Pseudoaffinity Chromatography Using a Convective Interaction Media Monolithic Column Hybridoma DOI: 10.1089/hyb.2011.0104

Dhivya AP, Kumar BP, Prasanna RR, Jayaprakash NS, & Vijayalakshmi MA (2010). Purification of Monoclonal Antibodies from Cell-Culture Supernatant by Use of Anion-Exchange Convective Interaction Media (CIM) Monolithic Columns Chromatographia, 72 (11-12) DOI: 10.1365/s10337-010-1787-3

Rediscovering the basics

Another paper on the purification of anti-HSA IgG1 MAb using CIM-IDA disks immobilized with four different metal ions. This time, the authors report superior purification with Zn (II) and Co (II) ions, with Zn (II)) being supposedly far better.

Again, similar problems as before: images with brightness and contrast adjusted to a point where the marker (especially in the "best" Zn (II) case) itself has lost some bands. In fact, the only ion with which they get one band is with Co (II) but its recovery does not appear to be very good.

Ironically in this paper, the authors make a statement (shown below) about how acidic pH is bad for their antibody. Given that the senior authors appear to have published many papers using pH 4 or pH 5.5 elution or equilibriation conditions (please see other posts in this blog), why this sudden discovery and what about those earlier antibodies and reports? 

Poonam Rajak et al. 2012                                                           Ambur P Dhivya et al. 2010

Fairly clean blot compared to other blots from this group with only slight degradation evident in the IgG1 MAb lane.

Rajak P, Vijayalakshmi MA, & Jayaprakash NS (2012). Purification of monoclonal antibodies, IgG1, from cell culture supernatant by use of metal chelate convective interaction media monolithic columns Biomedical Chromatography, 1488-1493 DOI: 10.1002/bmc.2721