In avian species, blood IgY is selectively incorporated into the yolks of maturing oocytes, although the precise mechanism is poorly understood. Our previous study showed that 22% of i.v.-injected heterologous chicken IgY (cIgY) was incorporated into egg yolks of Japanese quail (Coturnix japonica). However, it is not known whether homologous quail IgY (qIgY) can be more efficiently incorporated into quail egg yolks than cIgY. Therefore, we compared the uptakes of qIgY and cIgY i.v. administered into quail egg yolks and further characterized the uptakes of these 2 antibodies into quail ovarian follicles. Quail IgY and cIgY purified from the blood of the respective bird were labeled with digoxigenin, and their uptakes into quail egg yolks were determined by ELISA. Unexpectedly, total incorporation of the injected qIgY was only one-third of that of cIgY, although much more qIgY was left in blood compared with cIgY, suggesting that qIgY is the less preferable antibody as a transport ligand into quail egg yolks. On the other hand, deposition of the qIgY into heart, lung, liver, spleen, kidney, and ovarian follicular membrane was markedly higher than that of cIgY. Amino acid sequence analysis of 3 peptides derived from the trypsin-digested qIgY heavy chain revealed low homology between qIgY and cIgY. In conclusion, our results show that heterologous cIgY is more efficiently incorporated into quail egg yolks than homologous qIgY, possibly due to a distinctive antibody transport system existing in oocytes. The present results also may provide a new strategy for delivering useful proteinaceous substances into egg yolks in an attempt to produce designer eggs.

Maternal immunoglobulin (Ig) Y, the avian counterpart of mammalian IgG, is e ciently transported into the yolks of maturing oocytes. We have previously shown that the Fc region plays a critical role in the IgY transport into avian egg yolks. The aim of this study was to produce recombinant IgY-Fc and to evaluate its transport ability into avian egg yolks. Two basic expression vectors were constructed: one mainly expressed three constant regions of chicken IgY heavy chain (Fc) that contained three cysteine residues (C , C , C) involved in inter-heavy chain disulfide bonds; and the other mainly expressed two constant regions (Fc) that contained two cysteine residues (C , C). SDS-PAGE and Western blotting analyses indicated that both Fc and Fc were separated into multiple protein bands under non-reducing conditions, suggesting incomplete formation of inter-chain disulfide bonds. To prevent the incomplete disulfide bond formation, C and C were mutated individually to serine by site-directed mutagenesis. As expected, each of the mutated recombinant IgY-Fc produced a single protein band by SDS-PAGE analysis. When intravenously-injected into laying quail, the two mutants, Fc C S and Fc C S, were transported into the egg yolks at high levels, whereas the other two mutants, Fc C S and Fc C S, were transported into the egg yolks at lower levels. In conclusion, we have succeeded in producing a recombinant IgY-Fc retaining a high transport ability into the avian egg yolks; this protein will be provided as a useful tool for studying the mechanism of IgY transport into egg yolks. : egg yolk, IgY transport, inter-chain disulfide bond, quail, recombinant IgY-Fc

In avian species, it has been assumed that an Fc receptor in the ovarian follicles mediates immunoglobulin Y (IgY) transport into the yolk. However, no such receptor responsible for IgY has been identified to date. To examine potential IgY binding activity in the entire ovarian follicle, whole-mount sections of quail ovarian follicle were incubated with the Fc fragment of chicken IgY (cIgY). Whole-mount frozen sections of the second largest ovarian follicle were prepared, and then the sections were incubated with digoxigenin-labeled Fc or Fab fragments of cIgY. Microscopic observation revealed that incubation with the cIgY-Fc fragment produced a binding signal in the inner layer of the ovarian follicular tissues, most likely in the granulosa cell layer. However, no such signal was detected when the sections were incubated with cIgY-Fab. Coincubation of the ovarian sections with Alexa488-labeled cIgY-Fc and antiserum raised against ZP1, an envelope protein specifically localized in the perivitelline layer, demonstrated that the source of the Fc binding signals partly coincided with the perivitelline layer. In conclusion, our data suggest that potential IgY binding substances interacting with the Fc domain are present in the inner layers of ovarian follicular tissues, most likely in the granulosa cell layer and/or in the perivitelline layer.

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This study investigated the effects of dietary supplementation of cheese byproduct on performance, egg quality and fatty acid profile of egg yolk lipids from laying hens. One hundred five 30-wk-old White leghorn laying hens were randomly distributed into five groups of twenty one hens each and maintained in individual laying cages for 4 weeks. The hens were assigned to five treatments that consisted of corn-soybean meal based diets containing 0, 1, 3, 5 or 10% of cheese byproduct. Feed intake and rate of egg production of hens were not significantly different across the treatments during the whole experiment (p>0.05). Similarly, egg yolk cholesterol level, egg weight, Haugh's unit, eggshell thickness, color, and strength were not significantly different across the treatments (p>0.05). The amount of C16:0 in egg yolk was not significantly different across the treatments, but that of C18:0 decreased with increased cheese byproduct (p<0.01). Monounsaturated fatty acid (C16:1 and C18:1) content in egg yolk was similar across the treatments. Total CLA and cis-9, trans-11 CLA content increased linearly with increased cheese byproduct (p<0.001), while trans-10, cis-12 CLA amount was not significantly different across the treatments (p>0.05). Total saturated fatty acid (SFA) in the egg yolk was decreased as the level of cheese byproduct including CLA increased (p<0.01). However, the amount of unsaturated fatty acids (UFA) such as monounsaturated fatty acids (MUFA), n-3 polyunsaturated fatty acids (PUFA), n-6 PUFA, and total PUFAs in the egg yolk were not significantly different across the treatments (p>0.05). Therefore, the present results showed that cheese byproduct beneficially improved the fatty acid composition of concern to human health in the egg yolk without adverse effects on egg quality.