The force-extension behavior of individual mitotic newt chromosomes was studied, using micropipette surgery and manipulation, for elongations up to 80 times native length. After elongations up to five times, chromosomes return to their native length. In this regime chromosomes have linear elasticity, requiring ϳ1 nN of force to be stretched to two times native length. After more than five times stretching, chromosomes are permanently elongated, with force hysteresis during relaxation. If a chromosome is repeatedly stretched to ϳ10 times native length and relaxed, a series of hysteresis loops are obtained that converge to a single reversible elastic response. For further elongations, the linear dependence of force on extension terminates at a force "plateau" of ϳ15-20 nN, near 30 times extension. After Ͼ30 times extensions, the elastic moduli of chromosomes can be reduced by more than 20-fold, and they appear as "ghosts": swollen, elongated, and with reduced optical contrast under both phase and differential interference contrast imaging. Antibody labeling indicates that histone proteins are not being lost during even extreme extensions. Results are interpreted in terms of extension and failure of chromatin-tethering elements; the force data allow estimates of the number and size of such connectors in a chromosome.

The bending rigidities of mitotic chromosomes isolated from cultured N. viridescens (newt) and Xenopus epithelial cells were measured by observing their spontaneous thermal bending fluctuations. When combined with simultaneous measurement of stretching elasticity, these measurements constrain models for higher order mitotic chromosome structure. We measured bending rigidities of B ϳ10 Ϫ22 N ⅐ m 2 for newt and ϳ10 Ϫ23 N ⅐ m 2 for Xenopus chromosomes extracted from cells. A similar bending rigidity was measured for newt chromosomes in vivo by observing bending fluctuations in metaphase-arrested cells. Following each bending rigidity measurement, a stretching (Young's) modulus of the same chromosome was measured in the range of 10 2 to 10 3 Pa for newt and Xenopus chromosomes. For each chromosome, these values of B and Y are consistent with those expected for a simple elastic rod, B Ϸ YR 4 , where R is the chromosome cross-section radius. Our measurements rule out the possibility that chromosome stretching and bending elasticity are principally due to a stiff central core region and are instead indicative of an internal structure, which is essentially homogeneous in its connectivity across the chromosome cross-section.

NMR investigation of volume-limited chemical and biological samples requires a radio frequency (RF) microcoil with high signal-to-noise ratio (SNR) over the region of interest. Conventional approaches using solenoidal microcoils provide high sensitivity (RF field strength per unit current) and spectral resolution but require the sample and sample container to be enclosed by the coil. Planar surface microcoils provide an alternative configuration that allows direct access to the sample, but with the sacrifice of RF field uniformity. In this study we evaluate a family of planar RF microcoils (500 MHz, Archimedean geometry, 1-6 turns, inner radius 0.75 to 4 mm, and conductor widths of 75, 100, and 200 m). The design, fabrication, and performance (electrical and NMR) of the coils are described. This coil configuration exhibits a high local SNR, easy fabrication, good electrical properties, and moderate spectral uniformity (suitable for imaging and low resolution NMR spectroscopy). This design can also be scaled to smaller dimensions. These results suggest that planar spiral RF microcoils will find applications in multimodality microscopy and microfludic devices where sample manipulation and coil integration with the microanalysis systems is necessary.

We report measurements of the dynamics of force relaxation in single mitotic chromosomes, following step strains applied with micropipettes of force constant approximately 1 nN/microm. The force relaxes exponentially after an elongation (l/l(0)) to less than 3x native length, with a relaxation time approximately 2 sec. This relaxation time corresponds to an effective viscosity approximately 10(5) times that of water. We experimentally rule out solvent flow into the chromosome as the mechanism for the relaxation time. Instead, the relaxation can be explained in terms of the disentanglement dynamics of approximately 80 kb chromatin loop domains.

ABSTRACT:We describe the use of planar submillimeter diameter radio frequency (RF) microcoils at 500 MHz (11.74 T) to image biologic samples, specifically late-stage frog oocytes (typical diameter 750 -1000 m) and isolated rat pancreatic islets (typical diameter 150 -200 m). For the frog oocyte we obtained a 25 ϫ 25 m in-plane resolution on a 100 m slice in 34 min, and for the pancreatic islets we obtained a 14 ϫ 14 m in-plane resolution on a 100-m slice in 68 min. These results demonstrate that it is possible to obtain images of living cells with high temporal and spatial resolution under conditions where viability in maintained and functional stimulation is possible. The extension of these studies to smaller diameter microcoils or arrays of microcoils may allow magnetic resonance imaging to be performed on individual cells in culture, and could lead to in vivo assessment of cell function within biocapsules or tissue implants.