Biomedical subjects

D E Laney

Publications and source records attributed to D E Laney.

6 recordsLinked to original sources

PubMed1998

Substructure and responses of cholinergic synaptic vesicles in the atomic force microscope.

The substructure and responses of individual 100-nm synaptic vesicles to osmotic stress have been probed with an atomic force microscope (AFM) operating in tapping mode. Cholinergic synaptic vesicles from the electric organ of Torpedo californica were imaged continuously as the osmolarity of the buffer was decreased. Vesicles in hyposmotic buffer lysed to form flat circular structures on the mica surface with a diameter about two times that of intact vesicles and a thickness of 7.2 +/- 1.7 nm, which can accommodate the lipid bilayer plus the internal proteoglycan. Images of intact vesicles in air reveal creases in the membrane surface. Phase mode AFM images of lysed vesicles in air show the presence of a material not seen on intact vesicles that might be intravesicular proteoglycan released from the membrane at very low osmotic and ionic strength.

Acetylcholine↗

PubMed1997

Properties of biomolecules measured from atomic force microscope images: a review.

AFM images can be used to obtain quantitative or qualitative information about the properties of biomaterials. Examples presented here are: (1) Persistence length measurements of moving and stationary DNA molecules. (2) Force mapping to measure properties such as the elasticity of cells and vesicles. (3) Phase mode imaging to detect variations in materials and properties of the sample surface. (4) Imaging of surfaces at different constant forces.

Animals↗

PubMed1997

Changes in the elastic properties of cholinergic synaptic vesicles as measured by atomic force microscopy.

Cholinergic synaptic vesicles from Torpedo californica have been probed with the atomic force microscope in aqueous buffers to map and measure their elastic properties. Elastic properties were mapped with a new atomic force microscope technique known as force mapping. Force mapping of vesicles showed that the centers of the vesicles are harder or stiffer than the peripheral areas in the three buffers that were investigated. These were an isoosmotic buffer, a hypoosmotic buffer, and an isoosmotic buffer with 5 mM CaCl2 added. The hardness of the vesicular centers was quantified by calculation of the elastic modulus. Elastic moduli were in the range of 2-13 x 10(5) Pa. Vesicular centers were hardest in calcium-containing buffer and softest in isoosmotic buffer. Hypotheses are presented for the composition and function of the hard centers.

Acetylcholine↗

PubMed1996

Atomic force microscopy of long and short double-stranded, single-stranded and triple-stranded nucleic acids.

Atomic force microscopy (AFM, also called scanning force microscopy) is proving to be a useful technique for imaging DNA. Thus it is important to push the limits of AFM imaging in order to explore both what types of DNA can be reliably imaged and identified and also what substrates and methods of sample preparation are suitable. The following advances in AFM of DNA are presented here. (i) DNA molecules as short as 25 bases can be seen by AFM. The short single-stranded DNAs imaged here (25 and 50 bases long) appeared globular in the AFM, perhaps because they are all capable of intramolecular base pairing and because the DNAs were in a Mg(ll) buffer, which facilitates intramolecular cross-bridging. (ii) AFM images in air of short double-stranded DNA molecules, 100-200 bp, gave lengths consistent with A-DNA. (iii) AFM images of poly (A) show both short bent lumpy molecules with an apparent persistence length of 40 nm and long straight molecules with an apparent persistence length of 600 nm. For comparison, the apparent persistence length for double-stranded DNA from phX-174 under the same conditions was 80 nm. (iv) Structures believed to be triple- stranded DNA were seen in samples of poly(dA.poly(dT) and poly (dG).poly(dC). These structures were twice as high as double-stranded DNA and the same width. (v) Entire molecules of lambda DNA, approx. 16 micron long, were imaged clearly in overlapping scans. (vi) Plasmid DNA was imaged on oxidized silicon, although less clearly than on mica.

Base Sequence↗

PubMed1996

DNA binding to mica correlates with cationic radius: assay by atomic force microscopy.

In buffers containing selected transition metal salts, DNA binds to mica tightly enough to be directly imaged in the buffer in the atomic force microscope (AFM, also known as scanning force microscope). The binding of DNA to mica, as measured by AFM-imaging, is correlated with the radius of the transition metal cation. The transition metal cations that effectively bind DNA to mica are Ni(II), Co(II), and Zn(II), which have ionic radii from 0.69 to 0.74 A. In Mn(II), ionic radius 0.82 A, DNA binds weakly to mica. In Cd(II) and Hg(II), respective ionic radii of 0.97 and 1.1 A, DNA does not bind to mica well enough to be imaged with the AFM. These results may to relate to how large a cation can fit into the cavities above the recessed hydroxyl groups in the mica lattice, although hypotheses based on hydrated ionic radii cannot be ruled out. The dependence of DNA binding on the concentrations of the cations Ni(II), Co(II), or Zn(II) shows maximal DNA binding at approximately 1-mM cation. Mg(II) does not bind DNA tightly enough to mica for AFM imaging. Mg(II) is a Group 2 cation with an ionic radius similar to that of Ni(II). Ni(II), Co(II), and Zn(II) have anomalously high enthalpies of hydration that may relate to their ability to bind DNA to mica. This AFM assay for DNA binding to mica has potential applications for assaying the binding of other polymers to mica and other flat surfaces.

Adsorption↗

PubMed1995

Applications for atomic force microscopy of DNA.

Tapping mode atomic force microscopy (AFM) of DNA in propanol, dry helium, and aqueous buffer each have specific applications. Resolution is best in propanol, which precipitates and immobilizes the DNA and provides a fluid imaging environment where adhesive forces are minimized. Resolution on exceptional images of DNA appears to be approximately 2 nm, sufficient to see helix turns in detail, but the smallest substructures typically seen on DNA in propanol are approximately 6-10 nm in size. Tapping AFM in dry helium provides a convenient way of imaging such things as conformations of DNA molecules and positions of proteins on DNA. Images of single-stranded DNA and RecA-DNA complexes are presented. In aqueous buffer DNA molecules as small as 300 bp have been imaged even when in motion. Images are presented of the changes in shape and position of circular plasmid DNA molecules.

Bacteriophage lambda↗