Coordination Chemistry of Uranyl Ions with Surface-Immobilized Peptides: An XPS Study.

The coordination chemistry of uranyl ions with surface immobilized peptides was studied using X-ray photoemission spectroscopy (XPS). All the peptides in the study were modified using a six-carbon alkanethiol as a linker on a gold substrate with methylene blue as the redox label. The X-ray photoemission spectra reveal that each modified peptide interacts differently with the uranyl ion. For all the modified peptides, the XPS spectra were taken in both the absence and presence of the uranium, and their comparison reveals that the interaction depends on the chemical group present in the peptides. The XPS results show that, among all the modified peptides in the current study, the (arginine)9 (R9) modified peptide showed the largest response to uranium. In the order of response to uranium, the second largest response was shown by the modified (arginine)6 (R6) peptide followed by the modified (lysine)6 (K6) peptide. Other modified peptides, (alanine)6 (A6), (glutamic acid)6 (E6) and (serine)6 (S6), did not show any response to uranium.

Controlling the Biological Behaviors of Polymer-Coated Upconverting Nanoparticles by Adjusting the Linker Length of Estrone Ligands.

The estrone ligand is used for modifying nanoparticle surfaces to improve their targeting effect on cancer cell lines. However, to date, there is no common agreement on the ideal linker length to be used for the optimum targeting performance. In this study, we aimed to investigate the impact of poly(poly ethylene glycol methyl ether methacrylate) (PPEGMEMA) linker length on the cellular uptake behavior of polymer-coated upconverting nanoparticles (UCNPs). Different triblock terpolymers, poly(poly (ethylene glycol) methyl ether methacrylate)-block-polymethacrylic acid-block-polyethylene glycol methacrylate phosphate (PPEGMEMAx-b-PMAAy-b-PEGMP3: x = 7, 15, 33, and 80; y = 16, 20, 18, and 18), were synthesized with different polymer linker chain lengths between the surface and the targeting ligand by reversible addition-fragmentation chain transfer polymerization. The estrone ligand was attached to the polymer via specific terminal conjugation. The cellular association of polymer-coated UCNPs with linker chain lengths was evaluated in MCF-7 cells by flow cytometry. Our results showed that the bioactivity of ligand modification is dependent on the length of the polymer linker. The shortest polymer PPEGMEMA7-b-PMAA16-b-PEGMP3 with estrone at the end of the polymer chain was found to have the best cellular association behavior in the estrogen receptor (ER)α-positive expression cell line MCF-7. Additionally, the anticancer drug doxorubicin•HCl was encapsulated in the nanocarrier to evaluate the 2D and 3D cytotoxicity. The results showed that estrone modification could efficiently improve the cellular uptake in ERα-positive expression cell lines and in 3D spheroid models.

Evolution of an Electronic Health Record Based-Human Immunodeficiency Virus (HIV) Screening Program in an Urban Emergency Department for Diagnosing Acute and Chronic HIV Infection.

BACKGROUND: Since 2006, Centers for Disease Control and Prevention guidelines recommend routine opt-out human immunodeficiency virus (HIV) testing among sexually active 13- to 64-year-olds. Earlier diagnosis and treatment of HIV infection reduces morbidity and mortality and can limit transmission to others. OBJECTIVE: Our aim was to increase HIV testing, diagnosis, and linkage to care in the emergency department (ED). METHODS: Beginning May 4, 2015, we utilized our electronic health record (EHR) to enhance HIV testing in patients seen in the Rush University Medical Center emergency department in Chicago, IL, who were 13-64 years of age, did not have HIV listed on their problem list, and did not have an HIV antigen/antibody (Ag/Ab) test in the EHR within the past rolling 12-month period. Strategies included use of a "Best Practice Advisory" and later auto-order screening linked to a complete blood count order. RESULTS: Our baseline HIV test rate was 2.5% of the target population by age (average of 93 tests per month). From May 4, 2015 to January 31, 2019, 137,749 patients of 240,091 ED visits met our test criteria and 23,588 (17.1% of the target population) HIV Ag/Ab tests were performed, resulting in 164 positive tests. We identified 18 acute seroconverters, 51 new chronically infected persons, and 95 known infected, many of who had not disclosed their status. Our positive test rate was 0.70%, which dropped to 0.29% if only newly diagnosed individuals were counted. CONCLUSIONS: EHR enhancements in a large urban ED identifies both newly diagnosed acute and chronically HIV-infected persons. Identification of previously diagnosed patients offers an opportunity to relink them to care.

Effects of redox label location on the performance of an electrochemical aptamer-based tumor necrosis factor-alpha sensor.

We report the development of an electrochemical aptamer-based sensor for real time detection of tumor necrosis factor-alpha. The focus of this study is to evaluate the effects of the redox label location on the overall sensor performance, including sensor stability, detection limit, reusability, and selectivity. Three aptamer probes, each labeled with methylene blue (MB) at a specific location, were designed and employed in the fabrication of the sensors. Among the three sensors, the sensor fabricated using an aptamer with the MB label located at the distal end has a detection limit of 100 pM and is regenerable. The sensor fabricated using an aptamer with an internal MB modification has a detection limit of 10 nM and is not regenerable. Both sensors can be employed in complex biological samples such as 50% urine and 50% saliva. However, the sensor fabricated with an aptamer with the MB label located at the proximal end suffers from poor reproducibility and is highly unstable, thus limiting its application as a sensor. On the bases of these results, placing the MB label at the distal end of the aptamer probe appears to be the most advantageous for this sensor design for it does not interfere with monolayer formation and target binding.

Application of Calcium-Binding Motif of E-Cadherin for Electrochemical Detection of Pb(II).

We report, for the first time, the use of a 13-amino-acid peptide sequence derived from the calcium-binding site of E-cadherin in the fabrication of an electrochemical peptide-based (E-PB) Pb(II) sensor. The sensing mechanism is analogous to that of previously developed E-PB sensors. Binding of Pb(II) rigidifies the surface-immobilized and methylene blue (MB)-modified peptide probe, thereby limiting the accessibility of the tethered MB to the electrode surface. This change in probe flexibility results in a reduction in the MB current that is dependent on the target concentration. The sensor behaves as a "signal-off" sensor in alternating current voltammetry and cyclic voltammetry, but it can behave as a "signal-on" sensor in differential pulse voltammetry when a longer pulse width is employed. It is capable of specific detection of Pb(II) and is selective enough to be employed in realistically complex samples such as diluted tap water, saliva, and urine samples. The detection is fast; signal saturation can be achieved in <60 s. The sensor can also be fabricated on gold-plated screen-printed carbon electrodes, electrode substrates that are ideal for cost-effective analysis of Pb(II) in real-world settings.

Tunable Signal-Off and Signal-On Electrochemical Cisplatin Sensor.

We report the first electrochemical cisplatin sensor fabricated with a thiolated and methylene blue (MB)-modified oligo-adenine (A)-guanine (G) DNA probe. Depending on the probe coverage, the sensor can behave as a signal-off or signal-on sensor. For the high-coverage sensor, formation of intrastrand Pt(II)-AG adducts rigidifies the oligo-AG probe, resulting in a concentration-dependent decrease in the MB signal. For the low-coverage sensor, the increase in probe-to-probe spacing enables binding of cisplatin via the intrastrand GNG motif (N = A), generating a bend in the probe which results in an increase in the MB current. Although both high-coverage signal-off and low-coverage signal-on sensors are capable of detecting cisplatin, the signal-on sensing mechanism is better suited for real time analysis of cisplatin. The low-coverage sensor has a lower limit of detection, wider optimal AC frequency range, and faster response time. It has high specificity for cisplatin and potentially other Pt(II) drugs and does not cross-react with satraplatin, a Pt(IV) prodrug. It is also selective enough to be employed directly in 50% saliva and 50% urine. This detection strategy may offer a new approach for sensitive and real time analysis of cisplatin in clinical samples.

Electrochemical Detection of Platinum(IV) Prodrug Satraplatin in Serum.

We report the design and fabrication of a reagentless and reusable electrochemical sensor for detection of satraplatin (SAT), a platinum(IV) prodrug. The detection strategy is based on the electrocatalytic reaction between the Pt(IV) center of SAT and surface-immobilized methylene blue. We systematically evaluated the effect of passivating diluent chain length on the overall sensor performance. Our results show that the use of a shorter diluent like 2-mercaptoethanol is more advantageous than using a longer and more passivating diluent such as 6-mercapto-1-hexanol. Independent of the use of cyclic voltammetry or chronoamperometry as the sensor interrogation technique, all three sensors, each passivated with a different alkanethiol diluent, have been demonstrated to be sensitive; the limit of detection is in the range of 1-10 µM. They are also highly specific and do not respond to Pt(II) drugs such as cisplatin and carboplatin. More importantly, they are selective enough to be employed directly in 50% serum. This sensing strategy has potential applications in clinical pharmacokinetics studies.

Incorporation of extra amino acids in peptide recognition probe to improve specificity and selectivity of an electrochemical peptide-based sensor.

We investigated the effect of incorporating extra amino acids (AA) at the n-terminus of the thiolated and methylene blue-modified peptide probe on both specificity and selectivity of an electrochemical peptide-based (E-PB) HIV sensor. The addition of a flexible (SG)3 hexapeptide is, in particular, useful in improving sensor selectivity, whereas the addition of a highly hydrophilic (EK)3 hexapeptide has shown to be effective in enhancing sensor specificity. Overall, both E-PB sensors fabricated using peptide probes with the added AA (SG-EAA and EK-EAA) showed better specificity and selectivity, especially when compared to the sensor fabricated using a peptide probe without the extra AA (EAA). For example, the selectivity factor recorded in the 50% saliva was ∼2.5 for the EAA sensor, whereas the selectivity factor was 7.8 for both the SG-EAA and EK-EAA sensors. Other sensor properties such as the limit of detection and dynamic range were minimally affected by the addition of the six AA sequence. The limit of detection was 0.5 nM for the EAA sensor and 1 nM for both SG-EAA and EK-EAA sensors. The saturation target concentration was ∼200 nM for all three sensors. Unlike previously reported E-PB HIV sensors, the peptide probe functions as both the recognition element and antifouling passivating agent; this modification eliminates the need to include an additional antifouling diluent, which simplifies the sensor design and fabrication protocol.

Comparison of Mannose, Ethylene Glycol, and Methoxy-Terminated Diluents on Specificity and Selectivity of Electrochemical Peptide-Based Sensors.

We report the synthesis and application of three new antifouling diluents for the fabrication of an E-PB HIV sensor. Among the three thiolated antifouling diluents used in this study, the methoxy-terminated diluent (C6-MEG) is the most effective in alleviating both nonspecific binding and adsorption of matrix contaminants onto the sensor surface, especially when compared to the mannose- (C6-MAN) and ethylene-glycol-terminated (C6-EG) diluents. The sensor fabricated with C6-MEG has a specificity factor (∼13.5) substantially higher than the sensor passivated with only 6-mercapto-1-hexanol (∼1.5). It is functional even when employed directly in 25% serum, an achievement that has not been observed with this class of E-PB sensors. More importantly, incorporation of these antifouling diluents has negligible impact on other important sensor properties such as sensitivity and binding kinetics. This sensor passivation strategy is versatile and can potentially be used with other E-PB sensors, as well as surface-based sensors that utilize thiol-gold self-assembled monolayer chemistry.

A Hg(II)-mediated "signal-on" electrochemical glutathione sensor.

We report the design and fabrication of a DNA-based electrochemical sensor for detection of glutathione. Sensor signaling relies on glutathione's ability to chelate mercury Hg(II), displacing it from the thymine-Hg(II)-thymine complex formed between the surface-immobilized DNA probes. Our results show that this sensor is sensitive and selective enough to be employed in saliva.

An electrochemical peptide-based Ara h 2 antibody sensor fabricated on a nickel(II)-nitriloacetic acid self-assembled monolayer using a His-tagged peptide.

We report, for the first time, the use of a Ni(II)-nitriloacetic acid (NTA) self-assembled monolayer (SAM) in the fabrication of an electrochemical peptide-based (E-PB) sensor for detection of anti-Ara h 2 antibodies. We compared the performance of the sensor fabricated on a Ni(II)-NTA SAM using a His-tagged peptide with the sensor fabricated using the conventional approach via direct immobilization of a thiolated peptide. While both sensors responded only to the correct antibody in the presence of random antibodies, we observed differences between the sensors. Specifically, the detection limit of the His-tagged sensor was 1pM, significantly lower than the 200pM detection limit of the conventional thiolated sensor. More importantly, unlike our previously developed E-PB sensors, both sensors are regenerable and reusable. The thiolated sensor can be readily regenerated using guanidine hydrochloride; whereas the His-tagged sensor can be regenerated by direct displacement of the His-tagged probes using Ni(II) ions. Overall, our results show that both approaches are well-suited for E-PB sensor fabrication; more importantly, specific sensor properties such as detection limit and dynamic range can be tuned by simply using a different probe immobilization method.

Use of combined scanning electrochemical and fluorescence microscopy for detection of reactive oxygen species in prostate cancer cells.

Release of ROS from prostate cancer (PC3) cells was studied using scanning electrochemical microscopy (SECM) and fluorescence microscopy. One-directional lateral scan SECM was used as a rapid and reproducible tool for simultaneous mapping of cell topography and reactive oxygen species (ROS) release. Fluorescence microscopy was used in tandem to monitor the tip position, in addition to providing information on intracellular ROS content via the use of ROS-reactive fluorescent dyes. A unique tip current (iT) vs lateral distance profile was observed when the tip potential (ET) was set at -0.65 V. This profile reflects the combined effects of topographical change and ROS release at the PC3 cell surfaces. Differentiation between topographical-related and ROS-induced current change was achieved by comparing the scans collected at -0.65 and -0.85 V. The effects of other parameters such as tip to cell distance, solvent oxygen content, and scan direction on the profile of the scan were systematically evaluated. Cells treated with tert-butyl hydroperoxide, a known ROS stimulus, were also evaluated using the lateral scanning approach. Overall, the SECM results correlate well with the fluorescence results. The extracellular ROS level detected at the SECM tip was found to be similar to the intracellular ROS level monitored using fluorescence microscopy. While the concentration of each contributing ROS species has not been determined and is thus part of the future study, here we have successfully demonstrated the use of a simple two-potential lateral scan approach for analysis of ROS released by living cells under real physiological conditions.

Design and characterization of a metal ion-imidazole self-assembled monolayer for reversible immobilization of histidine-tagged peptides.

We report the design and characterization of a metal ion-imidazole self-assembled monolayer on a gold electrode. The resultant monolayer is well-suited for direct immobilization of histidine and methylene blue-modified peptides.

Design and characterization of an electrochemical peptide-based sensor fabricated via"click" chemistry.

We report an electrochemical peptide-based sensor fabricated via'click' chemistry for detection of anti-p24 antibodies. Our results also allude to a signaling mechanism similar to that of the linear probe electrochemical DNA sensor.

A folding-based electrochemical aptasensor for detection of vascular endothelial growth factor in human whole blood.

We herein report a folding-based electrochemical DNA aptasensor for the detection of vascular endothelial growth factor (VEGF) directly in complex biological samples, including blood serum and whole blood. The electrochemical signal generation is coupled to a large, target-induced conformational change in a methylene blue-modified and surface immobilized anti-VEGF aptamer. The sensor is sensitive, selective and essentially reagentless: we can readily detect VEGF down to 5 pM (190 pg/mL) directly in 50% blood serum. Similar to other aptasensors of this class, the VEGF sensor is also regenerable and reusable. In addition, the sensor performs comparably well even when fabricated on a gold-plated screen-printed carbon electrode and can potentially be implemented as a cost-effective, single-use biosensor for diseases diagnosis and therapy monitoring. The exceptional sensitivity, selectivity, and reusability of this electrochemical aptasensor platform suggest it may be a promising strategy for a wide variety of sensing applications.

Fabrication of an electrochemical DNA sensor array via potential-assisted "click" chemistry.

Here we report the fabrication of a 3-pixel electrochemical DNA sensor array via potential-assisted "click" chemistry. We found that the sensors in the array exhibit close to identical sensor performance when compared to sensors constructed via conventional "click" chemistry.

An electrochemical peptide-based biosensing platform for HIV detection.

We have fabricated a potentially generalizable electrochemical peptide-based (E-PB) sensor for the detection of HIV anti-p24 antibodies. The E-PB sensor is sensitive, specific and fares well even when challenged in a realistically complex medium such as human urine proxy.

Folding-based electrochemical DNA sensor fabricated on a gold-plated screen-printed carbon electrode.

Here we report a folding-based electrochemical DNA (E-DNA) sensor fabricated on a gold-plated screen-printed carbon electrode and show that the E-DNA sensor is not required to be fabricated on a relatively flat gold surface; the sensor works comparably well when fabricated on an electrodeposited gold film with a surface roughness factor of approximately 7.

A limited immunocytochemical panel for the distinction of subepithelial gastrointestinal mesenchymal neoplasms sampled by endoscopic ultrasound-guided fine-needle aspiration.

We studied the use of immunocytochemical analysis with material procured by endoscopic ultrasound-guided fine-needle aspiration (EUS-guided FNA) for the diagnosis of subepithelial intramural gastrointestinal (GI) mesenchymal neoplasms (SIGIMNs). We identified all EUS-guided FNA specimens of SIGIMNs that had undergone immunocytochemical analysis. Results were compared with follow-up histologic diagnoses. There were 95 aspirates that were diagnosed as GI mesenchymal tumors (GI stromal tumors [GISTs], n = 46), leiomyomas (n = 38), peripheral nerve sheath tumors (n = 5), and other neoplasms by cytologic examination. Immunoreactivity with antibodies to CD117 always predicted GIST at follow-up; 15 of 16 cases immunoreactive with antibodies to CD34 were found to be GISTs at follow-up. Strong immunoreactivity with antibodies to smooth muscle actin or desmin usually predicted a leiomyoma at follow-up aside from a single glomus tumor and a case with apparent nonneoplastic smooth muscle contaminant. When sufficient material is present, immunocytochemical analysis used with material obtained by EUS-guided FNA is highly predictive of final pathologic diagnosis.

Ciliated foregut cyst of pancreas: cytologic findings on endoscopic ultrasound-guided fine-needle aspiration.

The cytologic findings of a ciliated foregut cyst of the pancreas diagnosed by endoscopic ultrasound-guided fine-needle aspiration (FNA) are described. Cytologic features of ciliated foregut cysts include the presence of ciliated columnar cells and detached ciliary tufts in a cystic fluid background with amorphous debris and rare macrophages. These cytologic findings are clearly distinct from those of cystic mucinous neoplasms and other pancreatic cysts with which the ciliated foregut cyst may be confused. To the best of our knowledge, this is the first case reporting the cytologic findings of a pancreatic ciliated foregut cyst sampled by endoscopic ultrasound-guided FNA. We believe that the distinctive and characteristic cytologic features can allow a preoperative cytologic diagnosis of this highly unusual pancreatic cystic lesion.