Numerous fibrillar cords attaching IGC to HLB were not covered by gold nanoparticles marking coilin or snRNA (Supplementary Figs 4 and 5)

Numerous fibrillar cords attaching IGC to HLB were not covered by gold nanoparticles marking coilin or snRNA (Supplementary Figs 4 and 5). the surface topography and vice versa, nuclear body that do not share common molecular components may possess comparable topographical characteristics. We also have analyzed surface distribution of particular nuclear antigens (double stranded DNA, coilin and splicing snRNA) using indirect immunogold labeling with subsequent secondary electron detection of platinum Pyrroloquinoline quinone nanoparticles. We suggest that ultrastructural surface morphology reflects functional status of a nuclear body. Nuclear structures Pyrroloquinoline quinone are complex and dynamic parts of the cell that lack a surrounding membrane1,2. According to modern concept, nuclear non-membranous domains present as liquid droplets in the nucleoplasm with increased concentration of appropriate proteins and RNAs3,4. However, these domains possess unique boundaries separating them from the surroundings and preserving their integrity. Even though inner ultrastructural morphology and molecular composition of main conservative nuclear domains are well characterized, much less is known about ultrastructural topography of their surface. The main reason for the are the troubles of their isolation from your interphase nuclei. There are only a handful of works describing the ultrathin surface organization of the nuclear body in the interphase cells, namely biochemically isolated nucleolus5,6 and Pyrroloquinoline quinone Cajal body (CB)7. Surface topography of chromatin is quite well-characterized for the mammalian and herb metaphase chromosomes8,9,10 and for the polythene chromosomes from insect salivary glands11,12. Recent evidence suggest liquid-like structure not only for nuclear domains but also for chromatin13. Giant nuclei of CSF3R growing avian and amphibian oocytes represent a useful system for exploration of the surface ultrastructure of a number of nuclear body as well as transcriptionally active chromosomes. The main advantage here is the large size of the nucleus which allows manual isolation of intact nuclear structures into distinct preparations. Growing oocyte nucleus houses actively transcribed chromosomes in the lampbrush form and, in the case of amphibians, diverse extrachromosomal and chromosome associated nuclear body (numerous amplified nucleoli, histone locus body (HLB), interchromatin granule clusters (IGC) or B-snurposomes and other nuclear organelles)14,15,16,17,18. Recently, amplified nucleoli from Xenopus oocyte became a model object in defining the liquid-like behavior of nuclear body19. In contrast, the avian late-stage oocyte nucleus lacks nucleoli20,21,22 but contains chromosome-attached centromere protein body21,23,24. In some species of birds, Cajal body-like (CB-like) body were explained in the growing oocyte nucleus23. The inner ultrathin organization of the amphibian and avian oocyte nuclear structures has been Pyrroloquinoline quinone investigated in depth using transmission electron microscopy (TEM)23,25,26,27,28,29,30,31. Preparations of the oocyte nuclear content have been utilized for standard SEM as well. However, the ultrastructural surface topography was thoroughly explained only for the lampbrush chromosomes29,32,33,34. There are only few examples of the surface topography of nuclear body from amphibian oocytes analyzed using SEM32,35, and there is no published data on the surface topography of nuclear body from your avian oocytes. Finally, you will find no works analyzing the correlations between ultrathin topography and the distribution patterns of marker antigens on the surface of the oocyte nuclear structures. Low-voltage scanning electron microscopy (LVCSEM) is one of the techniques for analyzing surface topology of various biological samples with high resolution. Compared to standard SEM, LVCSEM with the secondary electron detection allows for analysis of the surface topography of uncoated, non-osmicated biological samples. Moreover, only in case of LVCSEM one has the opportunity to apply immunogold labelling techniques36,37. In this work we have aimed to visualize with high resolution the surface of microsurgically isolated nuclear body and giant lampbrush chromosomes from avian and amphibian oocytes. Our results demonstrate that LVCSEM without osmium fixation and conductive covering allows for the identification of extrachromosomal and chromosome-associated nuclear body in the oocyte nuclear content preparations. Moreover, individual lampbrush chromosome regions such as chromomeres, lateral loops with specific morphology of RNP-matrix, transcribed and untranscribed looped-out DNA regions are easily recognizable. Although usage of hypotonic treatments, chemical fixation, dehydration, and air-drying for samples preparation disturb liquid properties of nuclear structures, surface ultrastructure is usually properly preserved and generally displays.