Standard Chemistry Research Articles

Interference of Hemoglobin A1c Due to Hemoglobin Franklin Park

To the Editor: Hemoglobin (Hb)1 A1c (1) can be measured by cation-exchange HPLC, which separates hemoglobin fractions, including some Hb variants, based on the overall charge on the molecule. In populations for whom there is a high prevalence of homozygous sickle disease, it is advisable to routinely use to routinely use a separation technique such as HPLC because this allows identification of the HbS homozygous state (2). Sickle cell disease is marked by a shortened red cell life span, so clinical laboratories should not issue HbA1c results for cases of sickle disease. HbA1c immunoassays can be readily adapted to run on standard chemistry analyzers, but they …

Integrated Extreme Real-Time PCR and High-Speed Melting Analysis in 52 to 87 Seconds.

Extreme PCR in <30 s and high-speed melting of PCR products in <5 s are recent advances in the turnaround time of DNA analysis. Previously, these steps had been performed on different specialized instruments. Integration of both extreme PCR and high-speed melting with real-time fluorescence monitoring for detection and genotyping is presented here. A microfluidic platform was enhanced for speed using cycle times as fast as 1.05 s between 66.4 °C and 93.7 °C, with end point melting rates of 8 °C/s. Primer and polymerase concentrations were increased to allow short cycle times. Synthetic sequences were used to amplify fragments of hepatitis B virus (70 bp) and Clostridium difficile (83 bp) by real-time PCR and high-speed melting on the same instrument. A blinded genotyping study of 30 human genomic samples at F2 c.*97, F5 c.1601, MTHFR c.665, and MTHFR c.1286 was also performed. Standard rapid-cycle PCR chemistry did not produce any product when total cycling times were reduced to <1 min. However, efficient amplification was possible with increased primer (5 μmol/L) and polymerase (0.45 U/μL) concentrations. Infectious targets were amplified and identified in 52 to 71 s. Real-time PCR and genotyping of single-nucleotide variants from human DNA was achieved in 75 to 87 s and was 100% concordant to known genotypes. Extreme PCR with high-speed melting can be performed in about 1 min. The integration of extreme PCR and high-speed melting shows that future molecular assays at the point of care for identification, quantification, and variant typing are feasible.

The Deazidoalkoxylation: Sequential Nucleophilic Substitutions with Diazidated Diethyl Malonate.

Diazidated malonamides derived from amines and diazidated diethyl malonate react with lithiated alcohols through nucleophilic substitution reactions where azide acts as an unconventional leaving group. This deazidoalkoxylation leads to the formal construction of N, O-acetals, and the remaining azide functionality is a useful entry point for further functionalizations through, for example, standard cycloaddition chemistry. Thus, the presented chemistry provides an easy route toward densely functionalized molecules: Amines, alcohols, and alkynes can be attached onto the small malonate core unit in a sequential manner.

Photochemical Conjugation and One-Pot Radiolabelling of Antibodies for Immuno-PET.

Monoclonal antibodies (mAbs), immunoglobulin fragments, and other proteins are important scaffolds in the development of radiopharmaceuticals for diagnostic immuno-positron emission tomography (immuno-PET) and targeted radioimmunotherapy (RIT). Conventional methods for radiolabelling proteins with metal ions such as 68 Ga, 64 Cu, 89 Zr, and 90 Y require multi-step procedures involving pre-purification, functionalisation with a chelate, and subsequent radiolabelling. Standard coupling chemistries are time-consuming, difficult to automate, and involve synthesis, isolation, and storage of an intermediate, new molecular entity (the conjugated mAb) whose biochemical properties can differ from those of the parent protein. To circumvent these issues, we developed a photoradiochemical approach that uses fast, chemoselective, light-induced protein modification under mild conditions with novel metal-ion-binding chelates derivatised with aryl azide (ArN3 ) groups. Experiments show that one-pot photochemical conjugation and radiolabelling of formulated mAbs can be achieved in <20 min.

Preparation and Evaluation of ZD2 Peptide 64Cu-DOTA Conjugate as a Positron Emission Tomography Probe for Detection and Characterization of Prostate Cancer.

Positron emission tomography (PET) is a sensitive modality for cancer molecular imaging. We aim to develop a PET probe for sensitive detection and risk stratification of prostate cancer by targeting an abundant microenvironment oncoprotein, extradomain-B fibronectin (EDB-FN). The probe consists of a small ZD2 peptide specific to EDB-FN and a 64Cu-DOTA chelate. The probe was synthesized using standard solid-phase peptide chemistry and chelated to 64Cu prior to imaging. PET images were acquired at 4 and 22 h after intravenously injecting a 200 μCi probe into mice bearing human PC3 and LNCaP tumors, which represent highly aggressive and slow-growing prostate tumors, respectively. At 4 and 22 h postinjection, tumors could be clearly identified in the PET images. A significant higher signal was observed in PC3 tumors than in LNCaP tumors at 22 h (p = 0.01). Probe accumulation was also higher in PC3 tumors at 24 h. These data demonstrated that PET molecular imaging of EDB-FN in the tumor microenvironment of prostate cancer allows efficient differentiation of PC3 and LNCaP tumors in vivo. The ZD2 peptide-targeted PET probe shows potential in the detection and characterization of high-risk prostate cancer to improve the clinical management of prostate cancer.

Efficacy and safety of burosumab in children aged 1–4 years with X-linked hypophosphataemia: a multicentre, open-label, phase 2 trial

Children with X-linked hypophosphataemia have high concentrations of circulating phosphatonin fibroblast growth factor 23 (FGF23), which causes renal phosphate wasting and hypophosphataemia, rickets, skeletal deformities, and growth impairment. Burosumab, a human monoclonal antibody against FGF23, improves phosphate homoeostasis and rickets in children aged 5-12 years with X-linked hypophosphataemia. We aimed to assess the safety and efficacy of burosumab in younger children with X-linked hypophosphataemia. In this open-label, phase 2 trial at three hospitals in the USA, children (aged 1-4 years) with X-linked hypophosphataemia received burosumab (0·8 mg/kg) via subcutaneous injection every 2 weeks for 64 weeks. The dose was increased to 1·2 mg/kg if two consecutive pre-dose serum phosphorus concentrations were below 1·03 mmol/L (3·2 mg/dL), serum phosphorus had increased by less than 0·16 mmol/L (<0·5 mg/dL) from baseline, and a dose of burosumab had not been missed. Participants could continue to receive burosumab for up to an additional 96 weeks during the extension period. Key inclusion criteria were age 1-4 years at the time of informed consent; fasting serum phosphorus concentration of less than 0·97 mmol/L (3·0 mg/dL); serum creatinine 8·8-35·4 μmol/L (0·1-0·4 mg/dL); radiographic evidence of rickets (at least five participants were required to have a Thacher Rickets Severity Score of ≥1·5 at the knee); and a confirmed PHEX mutation or a variant of unknown significance in the patient or direct relative also affected with X-linked hypophosphataemia. Conventional therapy was stopped upon enrolment. The coprimary endpoints were safety and change from baseline to week 40 in fasting serum phosphorus concentrations. Changes in rickets severity from baseline to weeks 40 and 64 (assessed radiographically using Thacher Rickets Severity Score and an adaptation of the Radiographic Global Impression of Change), and recumbent length or standing height, were key secondary outcomes. This trial is registered with ClinicalTrials.gov, number NCT02750618, and is ongoing. Between May 16, 2016, and June 10, 2016, we enrolled 13 children with X-linked hypophosphataemia. All 13 children completed 64 weeks of treatment and were included in the efficacy and safety analysis; none exceeded 70 weeks of treatment at the time of analysis. Serum phosphorus least squares mean increase from baseline to week 40 of treatment was 0·31 mmol/L (SE 0·04; 95% CI 0·24-0·39; 0·96 mg/dL [SE 0·12]; p<0·0001). All patients had at least one adverse event. 14 treatment-related adverse events, mostly injection site reactions, occurred in five children. One serious adverse event considered unrelated to treatment (tooth abscess) occurred in a child with a history of tooth abscess. All other adverse events were mild to moderate, except a severe food allergy considered unrelated to treatment. No instances of nephrocalcinosis or noteworthy changes in the results of a standard safety chemistry panel emerged. Total Thacher Rickets Severity Score decreased by a least squares mean of -1·7 (SE 0·1; p<0·0001) from baseline to week 40 and by -2·0 (SE 0·1; p<0·0001) by week 64. The Radiographic Global Impression of Change score also indicated significant improvement, with a least squares mean score of +2·3 (SE 0·1) at week 40 and +2·2 (0·1) at week 64 (both p<0·0001). Mean length or standing height Z score was maintained from baseline to week 64. Burosumab had a favourable safety profile, increased serum phosphorus, and improved rickets and prevented early declines in growth in children aged 1-4 years with X-linked hypophosphataemia. These findings could substantially alter the treatment of young children with X-linked hypophosphataemia. Ultragenyx Pharmaceutical and Kyowa Kirin International.

Deep into the Water: Exploring the Hydro-Electromagnetic and Quantum-Electrodynamic Properties of Interfacial Water in Living Systems

Normal water structures are maintained largely by interactions with bi-omacromolecular surfaces and weak electromagnetic fields, which enable extended networks for electron and proton conductivity. All standard chemistry is totally reliant on electrostatics and avoids all mention of elec-trodynamics and the consequent radiation field, which is supporting the notion of water as a primary mediator of biological effects induced via electromagnetic means into living systems. Quantum Electrodynamic (QED) field theory have produced a vision of water in a liquid state as a medium, which for a peculiarity of its molecular electronic spectrum reveals itself as an essential tool for long-range communications, being able to change its supra-molecular organization in function of the interaction with the environment. This paper draws attention to the fact that interfacial water (nanoscale confined water) has been shown, independently by Emilio Del Giudice et al. and by Gerald Pollack et al., to contain respectively Coherence Domains (CDs) and Exclusion Zones (EZs), which may be regarded as long-range ensembles of CDs, dynamic aqueous structures, which uses the special properties of water, such as its electron/proton dynamics and organized response to electromagnetic fields, to receive electromagnetically encoded signals endowed with coherence (negentropy) at a low frequency, and sum the resultant excitations, so as to foster the redistribution of that coherence at frequencies which may affect biological systems. The phase transition of water from the ordinary coherence of its liquid state (bulk water) to the semi-crystalline or glassy and super-coherent state of interfacial water and its role in living organisms is discussed. The link between interfacial and intracellular water of the living and 1) the thermodynamic correlation between electron and proton transfer responsible for the redox potential of chemical species, 2) the Grotthuss mechanism and the H eightfold path, 3) superconductivity and superfluidity (dissipationless quantum states), 4) the proton motive force and protons role in biological liquid-flow systems, and 5) two possible explanations to as many non-ordinary phenomena, one related to the mind-body severely stressful condition due to a Near Death State (NDS) or to a Near Death Like State (NDLS), namely the Electromagnetic Hyper Sensitivity (EHS) or Electromagnetic After-Effect (EAE), the other related to the harmful consequences avoided during the so-called fire walking (ceremony), are discussed.

A general approach for the site-selective modification of native proteins, enabling the generation of stable and functional antibody-drug conjugates.

Antibody-drug conjugates (ADCs) are a class of targeted therapeutics that utilize the specificity of antibodies to selectively deliver highly potent cytotoxins to target cells. Although recent years have witnessed significant interest in ADCs, problems remain with the standard linkage chemistries used for cytotoxin-antibody bioconjugation. These typically (1) generate unstable constructs, which may lead to premature cytotoxin release, (2) often give a wide variance in drug-antibody ratios (DAR) and (3) have poor control of attachment location on the antibody, resulting in a variable pharmacokinetic profile. Herein, we report a novel divinylpyrimidine (DVP) linker platform for selective bioconjugation via covalent re-bridging of reduced disulfide bonds on native antibodies. Model studies using the non-engineered trastuzumab antibody validate the utility of this linker platform for the generic generation of highly plasma-stable and functional antibody constructs that incorporate variable biologically relevant payloads (including cytotoxins) in an efficient and site-selective manner with precise control over DAR. DVP linkers were also used to efficiently re-bridge both monomeric and dimeric protein systems, demonstrating their potential utility for general protein modification, protein stabilisation or the development of other protein-conjugate therapeutics.

Editors' Choice—Electric Field-Induced DNA Melting with Detection by Square Wave Voltammetry

Here we demonstrate a purely electrochemical approach for monitoring the electric field-induced melting of surface-bound DNA duplexes tethered to gold surfaces using standard surface attachment chemistry, i.e. single thiol-Au bonds. The sensitivity of square wave voltammetry is combined with the electric fields generated during programmed sequences of chronoamperometric pulses to enable a method for DNA analysis that can be carried out at room temperature without need for parallel spectroscopic monitoring. Electrochemical melting curves are obtained using both scanning potential pulses and constant potential pulses, which are analyzed to assess duplex stability and the extent of thiol desorption. The melting behavior is found to depend on the pulse potential and pule time. Under optimized conditions, thiol reduction is minimized and DNA duplexes can be discriminated based on the presence of a single base pair mismatch. The method is found to be less sensitive to the duplex length, presumably due to the rapid decay of the electric field away from the electrode surface. Based on these results, a simple model is proposed in which maintenance of the double-layer charge by accumulation of ions from the bulk of solution competes with electric field-induced loss of the negatively-charged DNA at a given applied potential.

Analysis of the Tailored Coupled-Cluster Method in Quantum Chemistry

In quantum chemistry, one of the most important challenges is the static correlation problem when solving the electronic Schrödinger equation for molecules in the Born--Oppenheimer approximation. In this article, we analyze the tailored coupled-cluster method (TCC), one particular and promising method for treating molecular electronic-structure problems with static correlation. The TCC method combines the single-reference coupled-cluster (CC) approach with an approximate reference calculation in a subspace (complete active space (CAS)) of the considered Hilbert space that covers the static correlation. A one-particle spectral gap assumption is introduced, separating the CAS from the remaining Hilbert space. This replaces the nonexisting or nearly nonexisting gap between the highest occupied molecular orbital and the lowest unoccupied molecular orbital usually encountered in standard single-reference quantum chemistry. The analysis covers, in particular, CC methods tailored by tensor-network states (TNS-TCC methods). The problem is formulated in a nonlinear functional analysis framework, and, under certain conditions such as the aforementioned gap, local uniqueness and existence are proved using Zarantonello's lemma. From the Aubin--Nitsche-duality method, a quadratic error bound valid for TNS-TCC methods is derived, e.g., for linear-tensor-network TCC schemes using the density matrix renormalization group method.

Combinatorial Peptide Microarray Synthesis Based on Microfluidic Impact Printing.

In this Research Article, a novel inkjet printing technique, micro impact printing (MI printing), is applied for the first time to combinatorial peptide microarray synthesis on amine functionalized microdisc arrays through standard Fmoc chemistry. MI printing shows great advantages in combinatorial peptide microarray synthesis compared with other printing techniques, including (1) a disposable cartridge; (2) a small spot size (80 μm) increases array density; (3) minimal loading volume (0.6 μL) and dead volume (<0.1 μL), reduce chemical waste; and (4) multiplexibility of 5 channels/cartridge and capacity of multiple cartridges. Using this synthesis platform, a tetrapeptide library with 625 permutations was constructed and then applied for the screening of ligands targeting α4β1 integrin on Jurkat cells.

Age-related phenotype and biomarker changes in SSADH deficiency.

ObjectiveSuccinic Semialdehyde Dehydrogenase (SSADH) deficiency is a disorder of elevated gamma‐amino butyric acid (GABA) and gamma hydroxybutyric acid (GHB) and a complex neuropsychiatric profile. Adult reports suggest worsening epilepsy and high SUDEP risk.MethodsSubjects with confirmed SSADH deficiency were recruited into a longitudinal study. Plasma thyroid hormone and total GABA/GHB were quantified by standard clinical chemistry methodologies and mass spectrometry, respectively.ResultsA total of 133 subjects with SSADH deficiency are enrolled in the registry; 49 participated in the longitudinal study. The age range of the population is 8 weeks to 63 years (median 7.75 year; 44% male). There is a significant difference in proportions among the age groups in subjects affected with hypotonia, compulsive behavior, sleep disturbances, and seizures. Epilepsy is present in 50% of the total population, and more prevalent in subjects 12 years and older (P = 0.001). The median age of onset for absence seizures was 2 years, and 12 years for generalized tonic‐clonic seizures (P < 0.01). The SUDEP rate in adults was 12% (4/33). There was a significant age‐dependent negative correlation between GABA and T3 levels.InterpretationThere is an age‐dependent association with worsening of epilepsy, behavioral disturbances including obsessive‐compulsive behavior, and sleep disturbances with age in SSADH deficiency. There is a high risk of SUDEP. We have observed more absence seizures in younger patients, compared to tonic‐clonic in the older cohort, which correlates with age‐related changes in GABA and GHB concentration and thyroid function, as well as the natural history of seizures in the murine model.

TEMPO-oxidized O-acetyl galactoglucomannan oligomers: isolation and comprehensive structural elucidation

Hemicelluloses, the second most abundant polysaccharides right after cellulose, represent complex biopolymers whose structural features are not yet fully understood. Hence, the applicability of functionalized hemicellulose derivatives is still limited. Thus, the development of practical methods to reveal their structure is essential. Anionic, carboxylated GGM was prepared as the substrate, and their corresponding oligomers have been isolated and fully characterized for the first time in the present work. In order to accomplish this, each step has been optimized sequentially. Oxidized GGM was enzymatically digested, and the corresponding oligomers were isolated by preparative GPC. A toolset of wet chemistry and instrumental methods was used to characterize the isolated compounds in detail. Moreover, quantitative 13C-NMR combined with MALDI was demonstrated to deliver accurate analytical data compared to standard but time-consuming wet chemistry methods. The developed toolset is expected to be applicable to other plant polysaccharides and their derivatives as well.

Synthesis of Fmoc-Protected ( S, S)- trans-Cyclopentane Diamine Monomers Enables the Preparation and Study of Conformationally Restricted Peptide Nucleic Acids.

An efficient synthesis of Fmoc-protected ( S, S)- trans-cyclopentane PNA ( tcypPNA) monomers starting from mono-Boc-protected ( S, S)-1,2-cyclopentanediamine is reported. A general synthetic strategy was developed so that tcypPNA monomers with each nucleobase can be made in sufficient quantity and purity for use in solid-phase peptide synthesis (SPPS). The newly synthesized monomers were then successfully incorporated into 10-residue PNA oligomers using standard Fmoc chemistry for SPPS. The different tcypPNAs allow different positions in the sequence to be conformationally constrained with ( S, S)- trans-cyclopentane to determine the effects on binding to complementary DNA.

Total Synthesis of Cyclic Octapeptide Reniochalistatin E

The total synthesis of a naturally occurring proline-enriched cyclic octapeptide, reniochalistatin E, from a marine sponge, where all the amino acids have the L-configuration, was reported. The reniochalistatin E was synthesized by applying a two-step solid-phase/solution synthesis strategy. The linear octapeptide was assembled by standard Fmoc chemistry on the solid phase, and subsequently cyclization was achieved by a solution method. The final product was purified by a preparative RP-HPLC system, and its structure was identified by HR-QTOF-MS, and 1H and 13C NMR.

Synthesis of Trinucleotide Building Blocks in Solution and on Solid Phase.

We have developed two methods, in solution and on solid phase, that give easy access to trinucleotide phosphoramidites capable of undergoing coupling reactions by the solid-phase phosphoramidite approach. The solution protocol is characterized by application of 5'-O-dimethoxytrityl (DMT) and 3'-O-tert-butyldimethylsilyl (TBDMS) as a pair of orthogonal protecting groups and 2-cyanoethyl (CE) for protection of the phosphate. Starting with suitably functionalized monomers, synthesis proceeds in the 3'- to 5'-direction, delivering the fully protected trinucleotide. The 3'-O-protecting group is cleaved followed by phosphitylation of the free 3'-OH group. The solid-phase protocol is based on standard phosphoramidite chemistry in conjunction with a dithiomethyl linkage connecting the 3'-starting nucleoside to the polymer. The disulfide bridge can be cleaved under neutral conditions for release of the trinucleotide from the support preserving all other protecting groups. © 2018 by John Wiley & Sons, Inc.

Characterization and detection of cardiac Troponin-T protein by using ‘aptamer’ mediated biofunctionalization of ZnO thin-film transistor

In this work, we have demonstrated the use of DNA aptamer as a selective bio recognizable entity towards the qualitative detection of cardiac Troponin-T (cTnT), a clinically important cardiac marker protein. Prior to this, the ZnO thin film surface as an active semiconducting channel material was first biofunctionalized with DNA aptamer by using standard biotin-streptavidin chemistry. Biofunctionalization was further confirmed by analyzing the ZnO surface with atomic force microscope (AFM) and X- ray photoelectron spectroscopy (XPS) techniques. In order to understand the efficacy of biomolecular interaction between cardiac Troponin-T (cTnT) and its corresponding aptamer, Kelvin Probe Force Microscopy (KPFM) technique was performed to monitor the effective contact potential difference. Further, to realize the essential technological importance of this methodology as a sensor platform, a thin-film transistor (TFT) micro device was fabricated using ZnO as a channel material and characterized. These fabricated devices were further used to demonstrate the qualitative detection of cardiac Troponin-T protein by employing aptamer-mediated biofunctionalization of ZnO TFT.

Compost Cation Exchange Capacity via Portable X-Ray Fluorescence (PXRF) Spectrometry

Compost is a valuable organic amendment which affords substantive fertility to soils where applied. A common component of compost fertility is cation exchange capacity (CEC), which has traditionally been determined via standard wet chemistry laboratory methods. This research utilized portable X-ray fluorescence (PXRF) spectrometry to evaluate 74 compost samples from the USA and Canada. PXRF elemental data were used for predicting compost CEC via random forest (RF) regression. Comparison between laboratory-determined vs. PXRF predicted CEC produced the following relationships: R2=0.90, RMSE = 5.41 meq 100 g−1 (model calibration) and R2=0.60, RMSE = 8.07 meq 100 g−1 (model validation). A key advantage of this technique is that the same data used for CEC prediction can also yield insight into other compost parameters of interest such as heavy metal content, plant essential nutrient content, salinity, and pH. Taken collectively, the PXRF approach can provide rapid, on-site analysis of compost which was previously not feasible with conventional methods. Our initial study has established the viability of PXRF for compost CEC determination, with further development on a wider array of feedstocks suggested for future study.

Novel Low Temperature Curable Photo-Patternable Polyamide with High Planarity for Wafer Level Fan-Out Packaging (WLFO)

Abstract As Wafer Level Fan-Out Packaging (WLFO) designs continue to evolve, higher pattern densities for Cu lines and interconnects continue to increase while thickness continues to decrease. As Cu densities increase, the patterning resolution of the RDL dielectric needs to shrink to allow increased bump density. The higher Cu density in turn requires enhanced dielectric performance to minimize Cu migration, whilst utilizing lower temperature and shorter cure times which result in lower levels of wafer warpage. Minimizing mechanical stress, continues to be a critical function of the RDL dielectric. Warpage leads to poor yields, distorting the planarity of the package and ultimately leading to stress induced failure from cracking and delamination. Reduction of the thermal budget is the primary means of reducing mechanical stress in WLFO designs. The amount of Cu in the package continues to increase. Differences in thermal expansions of such and the dielectric increase with temperature. Further, conventional solder reflow processes and set points will continue to be used, therefore the RDL dielectric must continue to be thermally and mechanically stable but capable of being cured at lower temperatures, 180–200 °C to minimize overall mechanical stresses from thermal expansion. We present a novel polyamide based RDL dielectric, KMRD, designed to achieve current and future WLFO design requirements. KMRD is a low temperature curable, aqueous (2.38%TMAH) developable dielectric capable of meeting industry standards for mechanical and electrical requirements, with a high level of reliability while utilizing a single stage cure at 185 °C. KMRD provides clear advantages over incumbent materials, with low temperature cure, improved pattern resolution, wide process latitudes, superior adhesion, chemical compatibility, and cost benefits using standard processing equipment and chemistry.

An Advanced, Silicon-Based Substrate for Sensitive Nucleic Acids Detection.

Surface substrate and chemical functionalization are crucial aspects for the fabrication of the sensitive biosensor based on microarray technology. In this paper, an advanced, silicon-based substrate (A-MA) allowing enhancement of optical signal for microarray application is described. The substrate consists in a multilayer of Si/Al/SiO2 layers. The optical signal enhancement is reached by a combination of the mirror effect of Al film and the SiO2 thickness around 830 nm, which is able to reach the maximum of interference for the emission wavelength of the Cy5 fluorescent label. Moreover, SiO2 layer is suitable for the immobilization of single-strand DNA through standard silane chemistry, and probe densities of about 2000 F/µm2 are reached. The microarray is investigated in the detection of HBV (Hepatitis B Virus) pathogen with analytical samples, resulting in a dynamic linear range of 0.05–0.5 nM, a sensitivity of about 18000 a.u. nM−1, and a Limit of Detection in the range of 0.031–0.043 Nm as a function of the capture probe sequence.