Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 4 de 4
Filter
Add more filters










Database
Language
Publication year range
1.
J Am Chem Soc ; 145(6): 3276-3282, 2023 02 15.
Article in English | MEDLINE | ID: mdl-36716175

ABSTRACT

For many classes of biomolecules, population-level heterogeneity is an essential aspect of biological function─from antibodies produced by the immune system to post-translationally modified proteins that regulate cellular processes. However, heterogeneity is difficult to fully characterize for multiple reasons: (i) single-molecule approaches are needed to avoid information lost by ensemble-level averaging, (ii) sufficient statistics must be gathered on both a per-molecule and per-population level, and (iii) a suitable analysis framework is required to make sense of a potentially limited number of intrinsically noisy measurements. Here, we introduce an approach that overcomes these difficulties by combining three techniques: a DNA nanoswitch construct to repeatedly interrogate the same molecule, a benchtop centrifuge force microscope (CFM) to obtain thousands of statistics in a highly parallel manner, and a Bayesian nonparametric (BNP) inference method to resolve separate subpopulations with distinct kinetics. We apply this approach to characterize commercially available antibodies and find that polyclonal antibody from rabbit serum is well-modeled by a mixture of three subpopulations. Our results show how combining a spatially and temporally multiplexed nanoswitch-CFM assay with BNP analysis can help resolve complex biomolecular interactions in heterogeneous samples.


Subject(s)
Antibodies , Nanotechnology , Animals , Humans , Rabbits , Bayes Theorem , Microscopy, Atomic Force/methods , Kinetics , Centrifugation/methods
2.
Nano Lett ; 21(1): 469-475, 2021 01 13.
Article in English | MEDLINE | ID: mdl-33395311

ABSTRACT

Molecular biomarkers play a key role in the clinic, aiding in diagnostics and prognostics, and in the research laboratory, contributing to our basic understanding of diseases. Detecting multiple and diverse molecular biomarkers within a single accessible assay would have great utility, providing a more comprehensive picture for clinical evaluation and research, but is a challenge with standard methods. Here, we report programmable DNA nanoswitches for multiplexed detection of up to 6 biomarkers at once with each combination of biomarkers producing a unique barcode signature among 64 possibilities. As a defining feature of our method, we show "mixed multiplexing" for simultaneous barcoded detection of different types of biomolecules, for example, DNA, RNA, antibody, and protein in a single assay. To demonstrate clinical potential, we show multiplexed detection of a prostate cancer biomarker panel in serum that includes two microRNA sequences and prostate specific antigen.


Subject(s)
DNA , MicroRNAs , Biomarkers, Tumor/genetics , DNA/genetics , MicroRNAs/genetics
3.
Sci Adv ; 5(3): eaau9443, 2019 03.
Article in English | MEDLINE | ID: mdl-30891499

ABSTRACT

MicroRNAs are short noncoding regulatory RNAs that are increasingly used as disease biomarkers. Detection of microRNAs can be arduous and expensive and often requires amplification, labeling, or radioactive probes. Here, we report a single-step, nonenzymatic microRNA detection assay using conformationally responsive DNA nanoswitches. Termed miRacles (microRNA-activated conditional looping of engineered switches), our assay has subattomole sensitivity and single-nucleotide specificity using an agarose gel electrophoresis readout. We detect cellular microRNAs from nanogram-scale RNA extracts of differentiating muscle cells and multiplex our detection for several microRNAs from one biological sample. We demonstrate 1-hour detection without expensive equipment or reagents, making this assay a compelling alternative to quantitative polymerase chain reaction and Northern blotting.


Subject(s)
DNA, Single-Stranded/metabolism , Electrophoresis, Agar Gel/methods , Genetic Engineering/methods , Inverted Repeat Sequences , MicroRNAs/analysis , Animals , Base Pairing , Cell Differentiation , Cell Line , DNA, Single-Stranded/genetics , Electrophoresis, Agar Gel/standards , Fluorescent Dyes/chemistry , Humans , Intercalating Agents/chemistry , Mice , MicroRNAs/genetics , MicroRNAs/metabolism , Myoblasts/cytology , Myoblasts/metabolism , Sensitivity and Specificity
4.
Nucleic Acids Res ; 45(19): 11459-11465, 2017 Nov 02.
Article in English | MEDLINE | ID: mdl-28977499

ABSTRACT

DNA serves as nature's information storage molecule, and has been the primary focus of engineered systems for biological computing and data storage. Here we combine recent efforts in DNA self-assembly and toehold-mediated strand displacement to develop a rewritable multi-bit DNA memory system. The system operates by encoding information in distinct and reversible conformations of a DNA nanoswitch and decoding by gel electrophoresis. We demonstrate a 5-bit system capable of writing, erasing, and rewriting binary representations of alphanumeric symbols, as well as compatibility with 'OR' and 'AND' logic operations. Our strategy is simple to implement, requiring only a single mixing step at room temperature for each operation and standard gel electrophoresis to read the data. We envision such systems could find use in covert product labeling and barcoding, as well as secure messaging and authentication when combined with previously developed encryption strategies. Ultimately, this type of memory has exciting potential in biomedical sciences as data storage can be coupled to sensing of biological molecules.


Subject(s)
Computers, Molecular , DNA, Viral/chemistry , Information Storage and Retrieval/methods , Nanostructures/chemistry , Bacteriophage M13/genetics , DNA, Viral/genetics , Electrophoresis, Agar Gel , Reproducibility of Results
SELECTION OF CITATIONS
SEARCH DETAIL
...