Patterns of mobilization of calcium,magnesium, and phosphorus by embryonic yellow-headed blackbirds(Xanthocephalus xanthocephalus)

Summary Embryonic blackbirds(Xanthocephalus xanthocephalus) obtain most of their calcium from the eggshell (85 90%), but all of their phosphorus comes from reserves in the yolk (80–85%) and albumen (15–20%). Approximately equal amounts of magnesium are supplied by the eggshell, the yolk, and the albumen. Yolk is depleted of magnesium and phosphorus during embryogenesis, but excess calcium absorbed from the eggeshell is stored in the yolk. Consequently reserves of calcium in the yolk actually increase 8-fold during embryonic development. Our results reveal that altricial birds manifest patterns of mobilization and deposition of calcium and other elements similar to those described for precocial species. Evolution of altriciality from precocity evidently did not entail major changes in how embryonic birds meet the challenge of obtaining the calcium, magnesium, and phosphorus required for development.     

乌骨鸡胚胎发育过程中卵内蛋白质脂肪的变化研究

对乌骨鸡胚胎发育过程中卵内各组成成分(蛋清、蛋黄、胚胎)在不同胚龄(即入孵的第0、3、6、9、12、15、18d和出壳)时蛋白质、脂肪含量变化进行测定与分析．结果表明：在不同胚龄时,蛋清中蛋白质含量显著高于蛋黄,蛋黄中脂肪含量始终高于蛋清;而胚胎中的蛋白质、脂肪逐渐增加。另外,胚胎发育过程中胚胎蛋白质的主要来源是蛋黄中的蛋白质,说明蛋黄是禽类胚胎发育过程中营养需要的主要来源。     

Egg Turning During Incubation has no Effect Upon the Growth of Embryos of Alligator mississippiensis

Alligator eggs are not turned during incubation, instead the embryo adheres to the top inside of the shell. Turning is alleged to shear off the embryo and kill it. Avian egg turning allegedly facilitates embryonic development by stimulating growth of the area vasculosa and minimizing the effects of unstirred yolk and albumen layers. From day 10 to day 45 of incubation, alligator eggs were experimentally turned, gently, through ± 60° in an hourly cycle. This turning regime killed only 6 out of 25 embryos. Compared with unturned controls, no significant effects were observed on the growth, production of extraembryonic fluids or utilization of albumen and yolk for those embryos that survived turning. The protein concentration of amniotic fluid at various stages of alligator development was examined in eggs incubated at 30 and 33°C. The fluid contained very little protein (max <8 mg) at any time: the protein concentration did not change consistently as development progressed. Differences in response to egg turning in birds and reptiles may be associated with the length of the incubation period, the protein content of the albumen and the mechanism of albumen utilization.     

Sources and timing of calcium mobilization during embryonic development of the corn snake, Pantherophis guttatus

Embryos of oviparous Reptilia (=turtles, lepidosaurs, crocodilians and birds) extract calcium for growth and development from reserves in the yolk and eggshell. Yolk provides most of the calcium to embryos of lizards and snakes. In contrast, the eggshell supplies most of the calcium for embryonic development of turtles, crocodilians and birds. The yolk sac and chorioallantoic membrane of birds recover and transport calcium from the yolk and eggshell and homologous membranes of squamates (lizards and snakes) probably transport calcium from these two sources as well. We studied calcium mobilization by embryos of the snake Pantherophis guttatus during the interval of greatest embryonic growth and found that the pattern of calcium transfer was similar to other snakes. Calcium recovery from the yolk is relatively low until the penultimate embryonic stage. Calcium removal from the eggshell begins during the same embryonic stage and total eggshell calcium drops in each of the final 2 weeks prior to hatching. The eggshell supplies 28% of the calcium of hatchlings. The timing of calcium transport from the yolk and eggshell is coincident with the timing of growth of the yolk sac and chorioallantoic membrane and expression of the calcium binding protein, calbindin-D28K, in these tissues as reported in previous studies. In the context of earlier work, our findings suggest that the timing and mechanism of calcium transport from the yolk sac of P. guttatus is similar to birds, but that both the timing and mechanism of calcium transport by the chorioallantoic membrane differs. Based on the coincident timing of eggshell calcium loss and embryonic calcium accumulation, we also conclude that recovery of eggshell calcium in P. guttatus is regulated by the embryo.     

Environmental modulation of calcium and phosphorus metabolism in embryonic snapping turtles (Chelydra serpentina)

Summary Eggs of common snapping turtles (Chelydra serpentina) were incubated on wet (–150 kPa water potential) and dry (–950 kPa) substrates in a laboratory study assessing the effects of the hydric environment on patterns of mobilization of calcium and phosphorus by developing embryos. We found that embryos developing in wet environments withdrew nutrients from their yolk faster, grew more rapidly, and incubated longer than embryos exposed to dry environments. Turtles developing in both environments absorbed calcium from the yolk at similar rates and depleted the yolk of almost its entire reserve of calcium prior to hatching. Calcium withdrawn from the yolk was supplemented with calcium mobilized from the eggshell, but embryos in wet environments obtained substantially more calcium from the eggshell than did those in dry settings. Embryos obtained all of the phosphorus used in skeletogenesis from the yolk, but those incubating in wet environments mobilized phosphorus from this compartment more rapidly than did those in dry settings. Exposing embryonic snapping turtles to wet environments apparently allows them to make more efficient use of the transitory source of calcium in the eggshell than is possible in dry environments. However, the residual yolk in hatchlings from both wet and dry environments contains too little calcium to support the growth of hard and soft tissues in neonates at rates similar to those characterizing the growth phase of development in embryogenesis.     

Effects of incubation temperature on growth and development of embryos ofAlligator mississippiensis

Summary Eggs ofAlligator mississippiensis were incubated at 30 °C and 33 °C throughout incubation up to hatching. Every four days several eggs were opened and the albumen, yolk and extra-embryonic fluids removed and weighed. The embryo was removed and fixed prior to being staged, weighted and measured for various morphometric criteria. Development at 33 °C was accelerated compared with 30 °C in terms of yolk and albumen utilization and embryo growth. Significant losses in yolk mass did not occur until stage 22 at 33 °C but occurred at stage 18 at 30 °C. Different patterns in growth were observed in embryos at the two temperatures at similar morphological stages: between stages 18 and 22 embryos at 33 °C were smaller (in mass and length) compared with embryos at 30 °C despite being morphologically similar. The differences in growth and physiology between embryos at 30 °C (females) and 33 °C (males) were dependent on incubation temperature but not sex. Incubation at 33 °C accelerated both growth and development inAlligator; initially morphogenesis was accelerated by the higher temperature but later, growth rate was accelerated.     

ACID-BASE RELATIONSHIPS WITHIN THE AVIAN EGG

1. In the newly laid egg of the domestic fowl the pH values of the albumen and yolk are about 7.6 and 6.0 respectively. 2. When the egg is stored in air there is a loss of carbon dioxide from the albumen and the pH of this fluid rises to a maximum value of about 9.5. A large proportion of the carbon dioxide which remains in the albumen is in the form of carbonate. 3. In the fertile incubated egg the pH of the albumen attains a maximum value within a period of about 2 days; the albumen then becomes less alkaline and it is nearly neutral by the end of the second week. The increasing acidity of the albumen can be attributed to (a) the secretion of hydrogen ions by the blastoderm and (b) the output of carbon dioxide by developing tissues. 4. During the first 2 weeks of incubation the pH of the yolk progressively increases to a maximum value of about 7.5: there is then a tendency for the pH of this fluid to fall and the yolk that is retained within the body of the hatched chick is slightly acidic. 5. The embryo may never come into direct contact with either the albumen or the yolk when the pH of these fluids are high and low respectively. At the beginning of embryonic development the blastoderm is separated from the albumen by the vitelline membrane and from the yolk by a layer of subgerminal fluid with a maximum pH of about 7.8. The vitelline membrane ruptures on day 4 but by this time the embryo is bathed in amniotic fluid with a pH of about 7.5. 6. The pH of amniotic fluid falls from a maximum value of about 7.5 during week I to a minimum value of about 6.5 during week 2. Amniotic fluid is a simple solution of salts until day 12; albumen then begins to flow into the amniotic cavity and the buffering capacity of amniotic fluid increases. 7. The principal end-product of nitrogenous metabolism in the chick embryo is uric acid and about 100 mg of this substance are deposited within the allantoic cavity. The pH of allantoic fluid may exceed 7.5 during week 1 but falls to 6.0 or below after day 13. 8. The tension of carbon dioxide within the egg is determined by the ratio of the rate of carbon dioxide production by the embryo to the permeability of the shell towards carbon dioxide. For the greater part of the period of incubation the permeability of the shell towards carbon dioxide is constant. Thus, as the carbon dioxide output of the embryo increases, the carbon dioxide tension within the egg rises. 9. The pH of the blood can be defined in terms of the ratio of the bicarbonate concentration to the carbon dioxide tension. There is a progressive increase in the carbon dioxide tension of the blood during the period of incubation but the pH is maintained at about 7.4 by an increase in bicarbonate concentration. 10. Part of the increase in bicarbonate is due to the removal of hydrogen ions from carbonic acid by haemoglobin. There is also a large influx of bicarbonate into the blood, but the source of this bicarbonate is not known; the evidence that renal mechanisms are involved is inconclusive and it is probable that the embryo utilizes the enormous potential store of bicarbonate in the egg shell.     

Growth and uptake of mineral by embryos of the direct-developing frog Eleutherodactylus coqui

Embryos of the direct-developing frog Eleutherodactylus coqui take up small quantities of yolk and yolk mineral early in incubation but increase their uptake of yolk reserves at later stages of development. Growth and accumulation of calcium and magnesium by embryos also occur slowly at first and at a higher rate later. Accumulation of calcium and magnesium by embryos is largely a function of variation in size of embryos, but uptake of phosphorus is unrelated to size. Althrough patterns of growth and uptake of mineral by embryonic coquis resemble those for embryos of oviparous amniotes, embryonic coquis do not deplete the yolk of its nutrients to the same degree. Thus, residual yolk of coqui hatchlings contains a high percentage of the nutrient reserves originally present in the egg. This difference between embryonic coquis and embryos of oviparous amniotes may indicate that transfer of nutrients from yolk to embryo becomes limiting during the grwoth phase. Alternatively, some aspects of the neurologic system are so poorly developed at hatching that coqui may not be able to find prey effectively. A large nutrient reserve could sustain hatchling while the neurologic system continues to mature.     

Selenium distribution in eggs of avian species

We studied the effect of egg mass of eight different avian species on Se distribution between egg components and the effect of incubation on Se accumulation by chicken eggshell and shell membrane. Eight groups of birds received a diet without Se supplementation. Unfertile eggs were collected after 35 days of feeding; yolk, albumen, shell and shell membrane were assayed separately for Se. All avian species studied showed identical Se concentration in yolk–albumen complex equal to 38.7 μg Se/100 g, reflecting a linear correlation between yolk–albumen mass and Se content. Shells and shell membrane Se accumulation showed quadratic correlation with the appropriate mass thus explaining unusually high Se concentration in ostrich shell and shell membrane, that reached values 1785 and 1904 μg Se/kg respectively. Incubation of fertile eggs decreased eggshell Se content, the effect being more expressed in eggs from hens fed sodium selenite compared to organic Se utilization (Sel-Plex). It was concluded that shell might be an additional Se source for an embryo.     

Effect of inorganic phosphate supplementation on egg production in Hy-Line Brown layers fed 2000 FTU/kg phytase

Phytase has long been used to decrease the inorganic phosphorus (Pi) input in poultry diet. The current study was conducted to investigate the effects of Pi supplementation on laying performance, egg quality and phosphate–calcium metabolism in Hy-Line Brown laying hens fed phytase. Layers (n = 504, 29 weeks old) were randomly assigned to seven treatments with six replicates of 12 birds. The corn–soybean meal-based diet contained 0.12% non-phytate phosphorus (nPP), 3.8% calcium, 2415 IU/kg vitamin D₃ and 2000 FTU/kg phytase. Inorganic phosphorus (in the form of mono-dicalcium phosphate) was added into the basal diet to construct seven experimental diets; the final dietary nPP levels were 0.12%, 0.17%, 0.22%, 0.27%, 0.32%, 0.37% and 0.42%. The feeding trial lasted 12 weeks (hens from 29 to 40 weeks of age). Laying performance (housed laying rate, egg weight, egg mass, daily feed intake and feed conversion ratio) was weekly calculated. Egg quality (egg shape index, shell strength, shell thickness, albumen height, yolk colour and Haugh units), serum parameters (calcium, phosphorus, parathyroid hormone, calcitonin and 1,25-dihydroxyvitamin D), tibia quality (breaking strength, and calcium, phosphorus and ash contents), intestinal gene expression (type IIb sodium-dependent phosphate cotransporter, NaPi-IIb) and phosphorus excretion were determined at the end of the trial. No differences were observed on laying performance, egg quality, serum parameters and tibia quality. Hens fed 0.17% nPP had increased (P < 0.01) duodenum NaPi-IIb expression compared to all other treatments. Phosphorus excretion linearly increased with an increase in dietary nPP (phosphorus excretion = 1.7916 × nPP + 0.2157; R² = 0.9609, P = 0.001). In conclusion, corn–soybean meal-based diets containing 0.12% nPP, 3.8% calcium, 2415 IU/kg vitamin D₃ and 2000 FTU/kg phytase would meet the requirements for egg production in Hy-Line Brown laying hens (29 to 40 weeks of age).     

Culture of naked quail (Coturnix coturnix japonica) ova in vitro for avian transgenesis: Culture from the single-cell stage to hatching with pH-adjusted chicken thick albumen

We first examined the pH change of the albumen of quail (Coturnix coturnix japonica) eggs before and after they were laid, as well as that of laid eggs. The pH rose rapidly after laying and continued to increase gradually in storage. Incubation at 37.5°C accelerated the increase in the pH of infertile eggs, while that of fertile eggs remained low during incubation. Referring to these results, we obtained a protocol for producing quail hatchlings by culture in vitro from naked ova. The naked ovum was filled with chicken (Gallus domesticus) thick albumen, the pH of which had been adjusted to 7.2–7.4. The ovum was cultured at 41.5°C in 20% CO₂ in air for the first 24 h. Then the embryo was moved to a surrogate quail egg shell that had been filled with non-pH-adjusted chicken thin albumen and cultured for a further 48 h in 100% air. The embryo was transferred again to a surrogate chicken egg shell and cultured under the same conditions until hatching. The culture yielded quail chicks with a hatchability of 19.4%. The method proposed here should be applicable to the production of transgenic birds.     

31P nuclear-magnetic-resonance studies on the developing embryos of Xenopus laevis.

The concentrations of nucleoside triphosphate, inorganic phosphate and the yolk proteins, phosvitin and lipovitellin, have been monitored in living embryos of Xenopus laevis by 31P nuclear magnetic resonance (NMR) spectroscopy. The nucleoside triphosphate levels remain relatively constant at about 3.5-4.5 nmol/embryo at least until the 'spontaneous movement' stage of development. By the swimming tadpole stage an inorganic phosphate resonance representing about 30 nmol/embryo becomes evident in the NMR spectrum. Computer manipulation also shows such a resonance, although smaller, to be present at a somewhat earlier developmental stage; these findings are confirmed biochemically. The major contribution to the NMR spectrum of oocytes, unfertilized eggs and early embryos is the yolk phosphoprotein resonance. On isolation of the yolk from the embryos it is possible to quantify the contribution to the NMR spectrum from the lipid-phosphate and protein-phosphate moieties of the yolk proteins. During development, as the yolk is used up, it is found that the protein-phosphate resonance disappears at a greater rate than the lipid-phosphate peak. The total phosphorus content of the embryo (approximately 200 nmol/embryo) is shown biochemically to remain constant during development; however, the total amount of phosphorus observed by NMR decreases by about 40% during development. From the resonance positions of their alpha, beta and gamma phosphate groups it is deduced that the nucleoside triphosphate molecules are liganded in vivo to a divalent cation which is not manganese, but could be either magnesium or calcium. From the position of the inorganic phosphate resonance it is deduced that the internal pH of embryos where this resonance is evident is 6.8 +/- 0.2.     

Localized axis determinant in the early cleavage embryo of the goldfish, Carassius auratus

 The teleost dorsoventral axis cannot be distinguished morphologically before gastrulation. In order to examine whether the yolk cell affects axis determination, we bisect early cleavage embryos of the goldfish, Carassius auratus. When the vegetal yolk hemisphere is removed by bisection along the equatorial plane at the 2-cell stage, the embryos develop abnormally and exhibit a symmetrical morphology. No dorsal structures, such as notochord, somites and neural tube, differentiate and no embryonic shield is formed during gastrulation. In addition, no goosecoid mRNA is expressed before gastrulation. The frequency of abnormality decreases as the age at which the vegetal yolk hemisphere is removed increases. Most embryos removed at the 32-cell stage develop normally. Their morphological phenotype is similar to that of a Xenopus ventralized embryo generated by ultraviolet irradiation on the vegetal hemisphere soon after fertilization. We also observed that, when the embryos were bisected along the first cleavage plane at the 2-cell stage, the proportion of pairs of embryos of which one embryo developed normally was 44.8%. These results indicate that the vegetal yolk hemisphere of the early cleavage embryo of the goldfish contains axis determination factor(s), which are necessary for generation of dorsal structures. Furthermore, it is suggested that these determinant(s) are distributed asymmetrically within the vegetal yolk hemisphere. Received: 25 May 1996 / Accepted: 19 September 1996     

Probable evolution of the coelacanth's reproductive style: lecithotrophy and orally feeding embryos in cichlid fishes and in Latimeria chalumnae

Synopsis The living coelacanth is a livebearer. Yolk seems to be the main source of nutrients and of oxygen to the embryo (fetus). Long before birth, young may also possibly feed orally on histotrophe secretion and egg debris. This type of reproduction evolved, as in most other fishes, from oviparity. The Carboniferous coelacanth Rhabdoderma exiguum had eggs of much lesser yolk volume and may represent an earlier form of oviparity with hiding, guarding or brooding type of parental care. The Jurassic coelacanth Holophagus (Undina) and the Cretaceous Axelrodichthys appear to have already evolved the internal-bearing style. Much of this evolutionary sequence is similar to that in cichlids. Ancestral cichlids are substrate tenders and nesters, with small eggs, little yolk and a feeding larva with indirect development. Mouthbrooding cichlids evolved a few, large eggs with denser yolk, direct development and, ultimately, orally feeding embryos while yolk is still in ample supply. Mixed feeding from yolk and orally ingested food in cichlids and in coelacanths is shown to be an enhanced mode of food delivery to the embryos over that from each source separately, in order to produce directly a better developed or larger young at the time of release, i.e. independence. Increase in egg size is regarded as an environmentally induced, altered pattern of yolk synthesis and an initial component of the epigenetic mechanism leading towards greater specialization. Carotenoids are incorporated within the yolk to assist the oxidative metabolism of the developing embryo.     

Embryonic modulation of maternal steroids in European starlings (Sturnus vulgaris)

In birds, maternally derived yolk steroids are a proposed mechanism by which females can adjust individual offspring phenotype to prevailing conditions. However, when interests of mother and offspring differ, parent–offspring conflict will arise and embryonic interests, not those of the mother, should drive offspring response to maternal steroids in eggs. Because of this potential conflict, we investigated the ability of developing bird embryos to process maternally derived yolk steroids. We examined how progesterone, testosterone and oestradiol levels changed in both the yolk/albumen (YA) and the embryo of European starling eggs during the first 10 days of development. Next, we injected tritiated testosterone into eggs at oviposition to characterize potential metabolic pathways during development. Ether extractions separated organic and aqueous metabolites in both the embryo and YA homogenate, after which major steroid metabolites were identified. Results indicate that the concentrations of all three steroids declined during development in the YA homogenate. Exogenous testosterone was primarily metabolized to an aqueous form of etiocholanolone that remained in the YA. These results clearly demonstrate that embryos can modulate their local steroid environment, setting up the potential for parent–offspring conflict. Embryonic regulation must be considered when addressing the evolutionary consequences of maternal steroids in eggs.     

Germline transmission of exogenous genes in chickens using helper-free ecotropic avian leukosis virus-based vectors

We have used vectors derived from avian leukosis viruses to transduce exogenous genes into early somatic stem cells of chicken embryos. The ecotropic helper cell line, Isolde, was used to generate stocks of NL-B vector carrying theNeo ^r selectable marker and theEscherichia coli lacZ gene. Microinjection of the NL-B vector directly beneath unincubated chicken embryo blastoderms resulted in infection of germline stem cells. One of the 16 male birds hatched (6.25%) from the injected embryos contained vector DNA sequences in its semen. Vector sequences were transmitted to G₁ progeny at a frequency of 2.7%.Neo ^r andlacZ genes were transcribedin vitro in chicken embryo fibroblast cultures from transgenic embryos of the G₂ progeny.     

The Effects of Experimentally Induced Adelphophagy in Gastropod Embryos

Adelphophagy, development where embryos grow large by consuming morphologically distinct nutritive embryos or their own normal siblings is widespread but uncommon among animal phyla. Among invertebrates it is particularly common in some families of marine gastropods and segmented worms, but rare or unknown in other closely related families. In calyptraeid gastropods phylogenetic analysis indicates that adelphophagy has arisen at least 9 times from species with planktotrophic larval development. This pattern of frequent parallel evolution of adelphophagy suggests that the embryos of planktotrophic species might be predisposed to evolve adelphophagy. Here we used embryos of three species of planktotrophic calyptraeids, one from each of three major genera in the family (Bostrycapulus, Crucibulum, and Crepidula), to answer the following 3 questions: (1) Can embryos of species with planktotrophic development benefit, in terms of pre-hatching growth, from the ingestion of yolk and tissue from experimentally damaged siblings? (2) Does ingestion of this material from damaged siblings increase variation in pre-hatching size? and (3) Does this experimentally induced adelphophagy alter the allometry between the velum and the shell, increasing morphological similarity to embryos of normally adelphophagic species? We found an overall increase in shell length and velum diameter when embryos feed on damaged siblings within their capsules. There was no detectable increase in variation in shell length or velum diameter, or changes in allometry. The overall effect of our treatment was small compared to the embryonic growth observed in naturally adelphophagic development. However each embryo in our experiment probably consumed less than one sibling on average, whereas natural adelphophages often each consume 10–30 or more siblings. These results suggest that the ability to consume, assimilate, and benefit from yolk and tissue of their siblings is widespread across calyptraeids.     

Avidin traps biotin diffusing out of chicken egg yolk

1. The unequal distribution of biotin and biotin-binding proteins between the yolk and albumen of freshly laid chicken eggs provides the potential for time-dependent redistribution of biotin that could affect egg quality, biotin availability, and hatchability. 2. Avidin-bound biotin was measured in albumen next to the shell and next to the yolk in eggs stored up to 23 days. 3. Biotin bound to biotin-binding proteins (BBP-I and BBP-II) was measured at the center and periphery of yolk from the same eggs. 4. After 11 days of storage, significant amounts of biotin from the yolk began to accumulate in the albumen adjacent to the yolk. 5. This transfer is attributed to a change in the vitelline membrane that permits diffusion of biotin, not BBP-I or BBP-II, out of the yolk. 6. The dynamics of this phenomenon suggest that in addition to its antimicrobial role, avidin may be involved in the utilization of biotin by the chick embryo.     

Effects of egg size on Double-crested Cormorant (Phalacrocorax auritus) egg composition and hatchling phenotype

Maternal investment of yolk and albumen in avian eggs varies with egg mass and contributes to variation in hatchling mass. Here we use the natural variation in mass and composition of Double-crested Cormorant (Phalacrocorax auritus) eggs to examine consequences of variation in yolk and albumen mass on hatchling phenotype. The Double-crested Cormorant, a large bird with altricial young, lays eggs ranging in mass from 40 to 60 g and containing an average of 82% albumen and 18% yolk. Variation in Cormorant egg mass arises primarily from variation in the amount of albumen and water in the eggs; yolk mass remains relatively constant, contributing only 10% to egg mass variation. Likewise, variation in hatchling mass correlates positively with albumen mass and albumen solids contribute to hatchling dry mass. Thus, variation in Cormorant egg mass is primarily the result of variation in the amount of egg albumen, which contributes most to variation in hatchling mass. Similarities in egg composition of altricial birds, along with data presented here, suggest that variation in hatchling mass of all altricial birds may depend most on the amount of egg albumen, unlike species with precocial young that hatch from eggs with substantially more yolk.     

Radioactive labeling of proteins in cultured postimplantation mouse embryos I. Influence of the embryo preparation method

Summary Conditions for optimum incorporation of radioactive amino acids into proteins of cultured postimplantation mouse embryos were investigated under the aspect of using these proteins for two-dimensional electrophoretic separations followed by fluorography. The aim was to obtain highly radioactive proteins under conditions as physiological as possible. Embryos at Days 10, 11, and 12 of gestation were prepared in different ways and incubated for 4 h in Tyrode’s solution containing [³H]amino acids (mixture) at a concentration of 27 μCi/ml medium. The preparations were: a) yolk sac opened, placenta and blood circulation intact; (b) yolk sac and amnion opened, placenta and blood circulation intact (Day 10 embryos only); c) placenta, yolk sac, and amnion removed (embryo “naked”); d) naked embryos cut randomly into pieces (Day 10 mebryos only). After incubation whole embryos or certain parts (tail, liver, rest body) were investigated by determining the radioactivity taken up by the protein. The results are given in dpm per mg protein per embryo. Radioactivity of proteins was about 3 times higher in naked mebryos than in embryos left in their yolk sacs. This was true for all three stages investigated. However, the degree of radioactivity in the various parts of naked embryos differed by a factor of 15, whereas radioactivity was evenly distributed in embryos incubated in their yolk sacs. Therefore, embryos prepared according to the first methods (see above) fulfilled the conditions required at the best. This work was supported by grants from the Deutsche Forschungsgemeinschaft awarded to the project K1 237/3-2 (Systematic Analysis of Cell Proteins).