Kovacs, P. | Search bioRxiv

bioRxiv2017

Widely used commercial ELISA for human Zonulin reacts with Complement C3 rather than pre-Haptoglobin 2

BACKGROUNDThere is increasing evidence for the role of impaired intestinal permeability in obesity and associated metabolic diseases. Zonulin is an established serum marker for intestinal permeability and identical to pre-haptoglobin2. Here, we aimed to investigate the relationship between circulating zonulin and metabolic traits related to obesity.\n\nMETHODSSerum zonulin was measured by using a widely used commercial ELISA kit in 376 subjects from the metabolically well-characterized cohort of Sorbs from Germany. In addition, haptoglobin genotype was determined in DNA samples from all study subjects.\n\nRESULTSAs zonulin concentrations did not correlate to the haptoglobin genotypes, we investigated the specificity of the zonulin ELISA assay using antibody capture experiments, mass spectrometry and Western blot analysis. Using serum samples that gave the highest or lowest ELISA signals, we detected several proteins that are likely to be captured by the antibody in the present kit. However, none of these proteins corresponds to pre-haptoglobin2. We used increasing concentrations of recombinant pre-haptoglobin 2 and complement C3 as one of the representative captured proteins and the ELISA kit did not detect either. Western blot analysis using both the polyclonal antibodies used in this kit and monoclonal antibodies rose against zonulin showed a similar protein recognition pattern but with different intensity of detection. The protein(s) measured using the ELISA kit was (were) significantly increased in patients with diabetes and obesity and correlated strongly with markers of the lipid and glucose metabolism. Combining mass spectrometry and Western blot analysis using the polyclonal antibodies used in the ELISA kit, we identified properdin as another member of the zonulin family.\n\nCONCLUSIONSOur study suggests that the zonulin ELISA does not recognize pre-haptoglobin 2, rather structural (and possibly functional) analogue proteins belonging to the mannose-associated serine protease family, with properdin being the most likely possible candidate.

molecular biology↗

bioRxiv2017

Face inversion reveals holistic processing of peripheral faces

Face perception is accomplished by face-selective neural processes, involving holistic processing that enables highly efficient integration of facial features into a whole face representation. It has been shown that in face-selective regions of the ventral temporal cortex, neural resources involved in holistic processing are primarily dedicated to the central portion of the visual field. These findings raise the intriguing possibility that holistic processing might be the privilege of centrally presented faces and could be strongly diminished in the case of peripheral faces. We addressed this question using the face inversion effect, a well established marker of holistic face processing. The behavioral results revealed impaired identity discrimination performance for inverted peripheral faces scaled according to the V1 magnification factor, compared to upright presented faces. The size of peripheral face inversion effect (FIE) was comparable to that found for centrally displayed faces. Face inversion affected the early ERP responses to faces in two time intervals. The earliest FIE was most pronounced in the time window between 130-140 ms following stimulus presentation, for both centrally and peripherally displayed faces and in the latter case, it was present only over the contralateral hemisphere. The timing of the next component FIE corresponded closely with the temporal interval of the N170 ERP component and showed strong right hemisphere lateralization, both when faces were displayed in the left or right visual field. Furthermore, we also showed that centrally presented face masks impaired peripheral face identity discrimination performance, but did not reduce the magnitude of the FIE. These findings revealed robust behavioral and neural inversion effects for peripheral faces and thus suggest that faces are processed holistically throughout the visual field.\n\nHighlightsRobust behavioral and neural inversion effect was found for peripheral faces.\n\nP1 ERP component is modulated by inverted central and contralateral faces.\n\nN170 ERP component is modulated by centrally and peripherally presented faces.\n\nNeural face inversion effect shows strong right hemisphere lateralization.\n\nFaces are processed holistically throughout the visual field.

neuroscience↗