Quantitative evaluation of oligonucleotide surface concentrations using polymerization-based amplification

Quantitative evaluation of minimal polynucleotide concentrations has become a critical analysis among a myriad of applications found in molecular diagnostic technology. Development of high-throughput, nonenzymatic assays that are sensitive, quantitative and yet feasible for point-of-care testing are thus beneficial for routine implementation. Here, we develop a nonenzymatic method for quantifying surface concentrations of labeled DNA targets by coupling regulated amounts of polymer growth to complementary biomolecular binding on array-based biochips. Polymer film thickness measurements in the 20–220 nm range vary logarithmically with labeled DNA surface concentrations over two orders of magnitude with a lower limit of quantitation at 60 molecules/μm2 (∼106 target molecules). In an effort to develop this amplification method towards compatibility with fluorescence-based methods of characterization, incorporation of fluorescent nanoparticles into the polymer films is also evaluated. The resulting gains in fluorescent signal enable quantification using detection instrumentation amenable to point-of-care settings

Sensitive Immunofluorescent Staining of Cells via Generation of Fluorescent Nanoscale Polymer Films in Response to Biorecognition

Immunofluorescent staining is central to nearly all cell-based research, yet only a few fluorescent signal amplification approaches for cell staining exist, each with distinct limitations. Here, the authors present a novel, fluorescent polymerization-based amplification (FPBA) method that is shown to enable similar signal intensities as the highly sensitive, enzyme-based tyramide signal amplification (TSA) approach. Being non-enzymatic, FPBA is not expected to suffer from nonspecific staining of endogenous enzymes, as occurs with enzyme-based approaches. FPBA employs probes labeled with photopolymerization initiators, which lead to the controlled formation of fluorescent polymer films only at targeted biorecognition sites. Nuclear pore complex proteins (NPCs; in membranes), vimentin (in filaments), and von Willebrand factor (in granules) were all successfully immunostained by FPBA. Also, FPBA was demonstrated to be capable of multicolor immunostaining of multiple antigens. To assess relative sensitivity, decreasing concentrations of anti-NPC antibody were used, indicating that both FPBA and TSA stained NPC down to a 1:100,000 dilution. Nonspecific, cytoplasmic signal resulting from NPC staining was found to be reduced up to 5.5-fold in FPBA as compared to TSA, demonstrating better signal localization with FPBA. FPBA’s unique approach affords a combination of preferred attributes, including high sensitivity and specificity not otherwise available with current techniques

DataSheet_5_Where the leatherbacks roam: movement behavior analyses reveal novel foraging locations along the Northwest Atlantic shelf.pdf

Leatherback sea turtles (Dermochelys coriacea) migrate along the east coast of the United States, traversing the South and Mid-Atlantic Bights (SAB and MAB) while traveling to and from well-known northern foraging areas off Southern New England (SNE) and Nova Scotia. However, there is limited information on leatherback movement behavior in these regions. To identify leatherback movement patterns, we fit hidden Markov models (HMMs) to satellite transmitter data from 52 leatherbacks tagged between 2017 and 2022 off the coasts of Massachusetts and North Carolina to estimate locations of area restricted searching (ARS) and transient behaviors. Depth-temperature profiles were then paired to locations associated with ARS behavior to understand the vertical use of the water column. We observed leatherbacks displaying ARS behavior in SNE as expected, but also in the MAB and SAB. The HMM results indicated that leatherbacks were primarily foraging in SNE between Nantucket and Long Island Sound and depth-temperature plots from ARS behavior on Nantucket Shoals implied turtles foraging throughout the entire water column. In the MAB, ARS behavior was concentrated between Cape Hatteras, North Carolina, and the mouth of Delaware Bay during the summer. Turtles were closely associated with a well-defined thermocline, but still appeared to dive to deeper cooler waters, which may be a sign of thermoregulatory behavior. There was evidence of foraging in the SAB along the coast as well as along the continental shelf edge. The ARS behavior we documented within the MAB and SAB is the first published empirical evidence that both areas may be important foraging grounds. Our results lay a path for future research to understand how leatherbacks use these areas and the potential anthropogenic threats encountered while moving through these regions.</p

DataSheet_7_Where the leatherbacks roam: movement behavior analyses reveal novel foraging locations along the Northwest Atlantic shelf.pdf

Leatherback sea turtles (Dermochelys coriacea) migrate along the east coast of the United States, traversing the South and Mid-Atlantic Bights (SAB and MAB) while traveling to and from well-known northern foraging areas off Southern New England (SNE) and Nova Scotia. However, there is limited information on leatherback movement behavior in these regions. To identify leatherback movement patterns, we fit hidden Markov models (HMMs) to satellite transmitter data from 52 leatherbacks tagged between 2017 and 2022 off the coasts of Massachusetts and North Carolina to estimate locations of area restricted searching (ARS) and transient behaviors. Depth-temperature profiles were then paired to locations associated with ARS behavior to understand the vertical use of the water column. We observed leatherbacks displaying ARS behavior in SNE as expected, but also in the MAB and SAB. The HMM results indicated that leatherbacks were primarily foraging in SNE between Nantucket and Long Island Sound and depth-temperature plots from ARS behavior on Nantucket Shoals implied turtles foraging throughout the entire water column. In the MAB, ARS behavior was concentrated between Cape Hatteras, North Carolina, and the mouth of Delaware Bay during the summer. Turtles were closely associated with a well-defined thermocline, but still appeared to dive to deeper cooler waters, which may be a sign of thermoregulatory behavior. There was evidence of foraging in the SAB along the coast as well as along the continental shelf edge. The ARS behavior we documented within the MAB and SAB is the first published empirical evidence that both areas may be important foraging grounds. Our results lay a path for future research to understand how leatherbacks use these areas and the potential anthropogenic threats encountered while moving through these regions.</p

DataSheet_1_Where the leatherbacks roam: movement behavior analyses reveal novel foraging locations along the Northwest Atlantic shelf.pdf

Leatherback sea turtles (Dermochelys coriacea) migrate along the east coast of the United States, traversing the South and Mid-Atlantic Bights (SAB and MAB) while traveling to and from well-known northern foraging areas off Southern New England (SNE) and Nova Scotia. However, there is limited information on leatherback movement behavior in these regions. To identify leatherback movement patterns, we fit hidden Markov models (HMMs) to satellite transmitter data from 52 leatherbacks tagged between 2017 and 2022 off the coasts of Massachusetts and North Carolina to estimate locations of area restricted searching (ARS) and transient behaviors. Depth-temperature profiles were then paired to locations associated with ARS behavior to understand the vertical use of the water column. We observed leatherbacks displaying ARS behavior in SNE as expected, but also in the MAB and SAB. The HMM results indicated that leatherbacks were primarily foraging in SNE between Nantucket and Long Island Sound and depth-temperature plots from ARS behavior on Nantucket Shoals implied turtles foraging throughout the entire water column. In the MAB, ARS behavior was concentrated between Cape Hatteras, North Carolina, and the mouth of Delaware Bay during the summer. Turtles were closely associated with a well-defined thermocline, but still appeared to dive to deeper cooler waters, which may be a sign of thermoregulatory behavior. There was evidence of foraging in the SAB along the coast as well as along the continental shelf edge. The ARS behavior we documented within the MAB and SAB is the first published empirical evidence that both areas may be important foraging grounds. Our results lay a path for future research to understand how leatherbacks use these areas and the potential anthropogenic threats encountered while moving through these regions.</p

DataSheet_2_Where the leatherbacks roam: movement behavior analyses reveal novel foraging locations along the Northwest Atlantic shelf.pdf

Leatherback sea turtles (Dermochelys coriacea) migrate along the east coast of the United States, traversing the South and Mid-Atlantic Bights (SAB and MAB) while traveling to and from well-known northern foraging areas off Southern New England (SNE) and Nova Scotia. However, there is limited information on leatherback movement behavior in these regions. To identify leatherback movement patterns, we fit hidden Markov models (HMMs) to satellite transmitter data from 52 leatherbacks tagged between 2017 and 2022 off the coasts of Massachusetts and North Carolina to estimate locations of area restricted searching (ARS) and transient behaviors. Depth-temperature profiles were then paired to locations associated with ARS behavior to understand the vertical use of the water column. We observed leatherbacks displaying ARS behavior in SNE as expected, but also in the MAB and SAB. The HMM results indicated that leatherbacks were primarily foraging in SNE between Nantucket and Long Island Sound and depth-temperature plots from ARS behavior on Nantucket Shoals implied turtles foraging throughout the entire water column. In the MAB, ARS behavior was concentrated between Cape Hatteras, North Carolina, and the mouth of Delaware Bay during the summer. Turtles were closely associated with a well-defined thermocline, but still appeared to dive to deeper cooler waters, which may be a sign of thermoregulatory behavior. There was evidence of foraging in the SAB along the coast as well as along the continental shelf edge. The ARS behavior we documented within the MAB and SAB is the first published empirical evidence that both areas may be important foraging grounds. Our results lay a path for future research to understand how leatherbacks use these areas and the potential anthropogenic threats encountered while moving through these regions.</p

DataSheet_4_Where the leatherbacks roam: movement behavior analyses reveal novel foraging locations along the Northwest Atlantic shelf.pdf

Leatherback sea turtles (Dermochelys coriacea) migrate along the east coast of the United States, traversing the South and Mid-Atlantic Bights (SAB and MAB) while traveling to and from well-known northern foraging areas off Southern New England (SNE) and Nova Scotia. However, there is limited information on leatherback movement behavior in these regions. To identify leatherback movement patterns, we fit hidden Markov models (HMMs) to satellite transmitter data from 52 leatherbacks tagged between 2017 and 2022 off the coasts of Massachusetts and North Carolina to estimate locations of area restricted searching (ARS) and transient behaviors. Depth-temperature profiles were then paired to locations associated with ARS behavior to understand the vertical use of the water column. We observed leatherbacks displaying ARS behavior in SNE as expected, but also in the MAB and SAB. The HMM results indicated that leatherbacks were primarily foraging in SNE between Nantucket and Long Island Sound and depth-temperature plots from ARS behavior on Nantucket Shoals implied turtles foraging throughout the entire water column. In the MAB, ARS behavior was concentrated between Cape Hatteras, North Carolina, and the mouth of Delaware Bay during the summer. Turtles were closely associated with a well-defined thermocline, but still appeared to dive to deeper cooler waters, which may be a sign of thermoregulatory behavior. There was evidence of foraging in the SAB along the coast as well as along the continental shelf edge. The ARS behavior we documented within the MAB and SAB is the first published empirical evidence that both areas may be important foraging grounds. Our results lay a path for future research to understand how leatherbacks use these areas and the potential anthropogenic threats encountered while moving through these regions.</p

DataSheet_6_Where the leatherbacks roam: movement behavior analyses reveal novel foraging locations along the Northwest Atlantic shelf.pdf

Leatherback sea turtles (Dermochelys coriacea) migrate along the east coast of the United States, traversing the South and Mid-Atlantic Bights (SAB and MAB) while traveling to and from well-known northern foraging areas off Southern New England (SNE) and Nova Scotia. However, there is limited information on leatherback movement behavior in these regions. To identify leatherback movement patterns, we fit hidden Markov models (HMMs) to satellite transmitter data from 52 leatherbacks tagged between 2017 and 2022 off the coasts of Massachusetts and North Carolina to estimate locations of area restricted searching (ARS) and transient behaviors. Depth-temperature profiles were then paired to locations associated with ARS behavior to understand the vertical use of the water column. We observed leatherbacks displaying ARS behavior in SNE as expected, but also in the MAB and SAB. The HMM results indicated that leatherbacks were primarily foraging in SNE between Nantucket and Long Island Sound and depth-temperature plots from ARS behavior on Nantucket Shoals implied turtles foraging throughout the entire water column. In the MAB, ARS behavior was concentrated between Cape Hatteras, North Carolina, and the mouth of Delaware Bay during the summer. Turtles were closely associated with a well-defined thermocline, but still appeared to dive to deeper cooler waters, which may be a sign of thermoregulatory behavior. There was evidence of foraging in the SAB along the coast as well as along the continental shelf edge. The ARS behavior we documented within the MAB and SAB is the first published empirical evidence that both areas may be important foraging grounds. Our results lay a path for future research to understand how leatherbacks use these areas and the potential anthropogenic threats encountered while moving through these regions.</p

DataSheet_3_Where the leatherbacks roam: movement behavior analyses reveal novel foraging locations along the Northwest Atlantic shelf.pdf

Leatherback sea turtles (Dermochelys coriacea) migrate along the east coast of the United States, traversing the South and Mid-Atlantic Bights (SAB and MAB) while traveling to and from well-known northern foraging areas off Southern New England (SNE) and Nova Scotia. However, there is limited information on leatherback movement behavior in these regions. To identify leatherback movement patterns, we fit hidden Markov models (HMMs) to satellite transmitter data from 52 leatherbacks tagged between 2017 and 2022 off the coasts of Massachusetts and North Carolina to estimate locations of area restricted searching (ARS) and transient behaviors. Depth-temperature profiles were then paired to locations associated with ARS behavior to understand the vertical use of the water column. We observed leatherbacks displaying ARS behavior in SNE as expected, but also in the MAB and SAB. The HMM results indicated that leatherbacks were primarily foraging in SNE between Nantucket and Long Island Sound and depth-temperature plots from ARS behavior on Nantucket Shoals implied turtles foraging throughout the entire water column. In the MAB, ARS behavior was concentrated between Cape Hatteras, North Carolina, and the mouth of Delaware Bay during the summer. Turtles were closely associated with a well-defined thermocline, but still appeared to dive to deeper cooler waters, which may be a sign of thermoregulatory behavior. There was evidence of foraging in the SAB along the coast as well as along the continental shelf edge. The ARS behavior we documented within the MAB and SAB is the first published empirical evidence that both areas may be important foraging grounds. Our results lay a path for future research to understand how leatherbacks use these areas and the potential anthropogenic threats encountered while moving through these regions.</p