At the onset of lay, the hen must provide for the continuous output of nutrients in the eggs. It is well known that the levels of some of the vitamins in the blood increase when the hen begins to lay. Common, Rutledge L k Bolton (1946) noted a marked rise in the serum riboflavin content of laying hens, which could also be evoked in the immature pullet by injection of oestradiol dipropionate. In their investigation, the content of serum carotenoids also rose after the oestrogen treatment. Hertz,

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... Dhyse & Tullner (1949) induced similarly a rise in the biotin level of plasma. In view of these observations, we wanted to know whether increased ovarian activity during egg production brought about any changes in the level of vitamin B" in the blood. Accordingly the vitamin B" content of cells and plasma was determined in pullets before and after they had begun to lay and in immature pullets and cockerels receiving oestradiol dipropionate. The blood of birds differs from that of mammals in having nucleated erythrocytes. Thus, there is the possibility that deoxyribosides (with growth activity for Lactobacillus Zeichmannii) are released from the nucleic acids in the erythrocytes during the preparation of extracts of the blood for microbiological assay of vitamin B". The blood of the domestic fowl also has the property of combining with large quantities of added vitamin B", and the vitamin B" naturally present in it is in the form of a protein complex not readily available to assay micro-organisms (Gregory & Holdsworth, 1959) . For these reasons, routine methods for measuring vitamin B" in mammalian blood could not be used, and different methods for extracting and measuring the vitamin in the blood of the domestic fowl were therefore investigated. The method finally chosen as being most suitable was then used to study the effect of onset of lay, or of administration of oestrogens to immature birds, on the vitamin B" levels in the blood plasma and cells. EXPERIMENTAL Procedure Expt I. Vitamin B" content of whole blood. Three different methods for extracting the vitamin B" activity from whole blood were investigated. Methods I , 2 and 3, described on p. 585, were each used to prepare extracts from a sample of pooled blood from 4-week-old pullets. The vitamin B" activity of the extracts, before and after treatment at https://www.cambridge.org/core/terms. https://doi.