FORMALIN DILUTIONS FOR FIXATION OF Centropyge aurantonotus EGGS

The use of fixatives applied to morphological studies is necessary for tissue preservation. Formalin is a less expensive methodology for specimen fixation, and buffered formalin is used for fixing fish eggs and larvae. We compared different formalin dilutions for fixing pelagic eggs of pygmy angelfish Centropyge aurantonotus.

Eight solutions of formaldehyde (P.A., 36.5-38%, Dinâmica Química Contemporânea LTDA ®, São Paulo - BR) were tested (Table 1). In buffered treatments, sodium phosphate was used. Eggs obtained from a captive pair of C. aurantonotus were incubated in 3-L recipients, in a 27ºC water bath, at a density of 0.1 egg mL-1. In blastula and gastrula developmental stages, the eggs were photographed and transferred to 1.5 mL conical microtubes, where the fixative was added. After 24h of fixation, samples were photographed again. The average egg diameter, coloration, chorion and embryo integrity were evaluated using DinoCapture 2.0 software. Differences in egg size before and after fixation were submitted to Student's t-test (p <0.05) and for comparison between treatments one-way ANOVA (p <0.05) was used.

All dilutions caused a significant increase in egg size, which varied between treatments (Table 1). Buffered formalin diluted in distilled water did not change egg color and resulted in good embryo fixation. The chorion and the oil drop did not undergo extreme changes after fixation, except for 4% buffered formalin diluted in seawater, which caused deformations in the oil drop. The buffer did not show good dilution in seawater, which may have impaired the preservation of the sample. The formalin treatments without buffer caused deformities in the embryos and egg color, except for 2% formalin diluted in seawater, which kept the embryo intact.

Sodium phosphate buffered formalin diluted in distilled water and 2% non-buffered formalin diluted in seawater maintained the integrity of the embryo and oil drop of C. aurantonotus eggs. However, hydration after fixation should be taken into account in morphometric studies. Due to the benefit of the low cost, formalin at these dilutions may serve as an alternative for fixing pelagic eggs of marine fishes. Studies comparing formalin fixation with other fixatives are necessary to determine the most efficient fixative for this type of egg, in addition to define a post fixation preservative.

Acknowledgments: CNPq and CAPES for their support in research funding and scholarships.