Direct and Label-Free Determination of Human Glycated Hemoglobin Levels Using Bacteriorhodopsin as the Biosensor Transducer

Glycated hemoglobin (HbA1c) levels are an important index for the diagnosis and long-term control of diabetes. This study is the first to use a direct and label-free photoelectric biosensor to determine HbA1c using bacteriorhodopsin-embedded purple membranes (PM) as a transducer. A biotinylated PM (b-PM) coated electrode that is layered with protein A-oriented antibodies against hemoglobin (Hb) readily captures non-glycated Hb (HbA0) and generates less photocurrent. The spectra of bacteriorhodopsin and Hb overlap so the photocurrent is reduced because of the partial absorption of the incident light by the captured Hb molecules. Two HbA0 and HbA1c aptasensors that are prepared by conjugating specific aptamers on b-PM coated electrodes single-step detect HbA0 and HbA1c in 15 min, without cross reactivity, with detection limits of ≤0.1 μg/mL and a dynamic range of 0.1–100 μg/mL. Both aptasensors exhibit high selectivity and long-term stability. For the clinical samples, HbA0 concentrations and HbA1c levels that are measured with aptasensors correlate well with total Hb concentrations and the HbA1c levels that are determined using standard methods (correlation gradient = 0.915 ± 0.004 and 0.981 ± 0.001, respectively). The use of these aptasensors for diabetes care is demonstrated.

Download Full-text

Multitarget, quantitative nanoplasmonic electrical field-enhanced resonating device (NE2RD) for diagnostics

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1510824112 ◽

2015 ◽

Vol 112 (32) ◽

pp. E4354-E4363 ◽

Cited By ~ 44

Author(s):

Fatih Inci ◽

Chiara Filippini ◽

Murat Baday ◽

Mehmet Ozgun Ozen ◽

Semih Calamak ◽

...

Keyword(s):

Dynamic Range ◽

Point Of Care ◽

Medical Diagnostics ◽

Clinical Samples ◽

Label Free ◽

Protein Biomarkers ◽

Electrical Field ◽

Color Changes ◽

Sensing Platform ◽

Broad Dynamic Range

Recent advances in biosensing technologies present great potential for medical diagnostics, thus improving clinical decisions. However, creating a label-free general sensing platform capable of detecting multiple biotargets in various clinical specimens over a wide dynamic range, without lengthy sample-processing steps, remains a considerable challenge. In practice, these barriers prevent broad applications in clinics and at patients’ homes. Here, we demonstrate the nanoplasmonic electrical field-enhanced resonating device (NE2RD), which addresses all these impediments on a single platform. The NE2RD employs an immunodetection assay to capture biotargets, and precisely measures spectral color changes by their wavelength and extinction intensity shifts in nanoparticles without prior sample labeling or preprocessing. We present through multiple examples, a label-free, quantitative, portable, multitarget platform by rapidly detecting various protein biomarkers, drugs, protein allergens, bacteria, eukaryotic cells, and distinct viruses. The linear dynamic range of NE2RD is five orders of magnitude broader than ELISA, with a sensitivity down to 400 fg/mL This range and sensitivity are achieved by self-assembling gold nanoparticles to generate hot spots on a 3D-oriented substrate for ultrasensitive measurements. We demonstrate that this precise platform handles multiple clinical samples such as whole blood, serum, and saliva without sample preprocessing under diverse conditions of temperature, pH, and ionic strength. The NE2RD’s broad dynamic range, detection limit, and portability integrated with a disposable fluidic chip have broad applications, potentially enabling the transition toward precision medicine at the point-of-care or primary care settings and at patients’ homes.

Download Full-text

P1958Baseline glycated hemoglobin level was differently associated with long-term prognosis in peripheral artery disease patients with and without hemodialysis

European Heart Journal ◽

10.1093/eurheartj/ehz748.0705 ◽

2019 ◽

Vol 40 (Supplement_1) ◽

Author(s):

S Shirotani ◽

K Jujo ◽

K Mizobuchi ◽

I Ishida ◽

Y Minami ◽

...

Keyword(s):

Peripheral Artery Disease ◽

Cox Regression ◽

Glycated Hemoglobin ◽

Baseline Hba1c ◽

Peripheral Artery ◽

Cox Regression Analysis ◽

Artery Disease ◽

Long Term Prognosis ◽

Hba1c Levels

Abstract Introduction Diabetes (DM) is the leading cause of induction of hemodialysis (HD), and both are major prognostic factors in patients with peripheral artery disease (PAD). However, the prognostic importance of baseline glycated hemoglobin (HbA1c) levels in PAD patients with and without HD is yet to be elucidated. Purpose We hypothesized that baseline HbA1c levels had different prognostic impacts after endovascular therapy (EVT) in between PAD patients with and those without regular HD. Methods This observational study included 643 consecutive patients who received EVT between 2013 and 2017. Each of 313 HD patients and 330 Non-HD patients was respectively divided into 2 groups by the comorbidity of diabetes, and DM patients were further divided into 3 subgroups depending on HbA1c level at the time of EVT; DM-Low (HbA1c: <6.0%), DM-Mid (6.1–7.0%), and DM-High (>7.1%) groups. The primary endpoint of this study was major amputation-free survival (AFS). Results HD group included significantly more patients presenting critical limb ischemia than Non-HD group (46.6% vs. 30.0%, p<0.001). During the observation period after EVT, there were 81 events (25.9%), including 55 death and 26 major amputations in HD group, and 45 events (13.6%), including 30 death and 15 major amputations in Non-HD group. Kaplan-Meier analysis revealed that DM patients had a significantly higher AFS rate in HD group (Log-rank: p=0.003, Figure A). In contrast, in Non-HD group, there was no statistical difference in AFS between DM and Non-DM patients (p=0.36). In Cox regression analysis in HD group, the higher HbA1c-DM group showed the higher hazard ratio (HR) for AFS (p=0.039, Figure B), and DM patients with HbA1c >6.0% had significantly higher HR than Non-DM patients. Whereas, in Non-HD group, HbA1c-DM class adversely associated with HR for AFS (p=0.003), even any classes did not reach statistical differences from Non-DM patients. Figure 1 Conclusions Comorbidity of diabetes at the time of EVT worsened long-term prognosis in PAD patients receiving regular HD, but not in those without HD. Additionally, baseline HbA1c levels oppositely affected prognosis in PAD patients with and without HD.

Download Full-text

Macroprolactinemia: Diagnostic, Clinical, and Pathogenic Significance

Clinical and Developmental Immunology ◽

10.1155/2012/167132 ◽

2012 ◽

Vol 2012 ◽

pp. 1-7 ◽

Cited By ~ 38

Author(s):

Akira Shimatsu ◽

Naoki Hattori

Keyword(s):

Middle Age ◽

Molecular Form ◽

Protein A ◽

Gel Filtration ◽

Cross Reactivity ◽

Precipitation Method ◽

Binding Studies ◽

Rat Pituitary ◽

Peg Precipitation Method

Macroprolactinemia is characterized by a large molecular mass of PRL (macroprolactin) as the main molecular form of PRL in sera, the frequent elevation of serum PRL (hyperprolactinemia), and the lack of symptoms. Macroprolactin is largely a complex of PRL with immunoglobulin G (IgG), especially anti-PRL autoantibodies. The prevalence of macroprolactinemia is 10–25% in patients with hyperprolactinemia and 3.7% in general population. There is no gender difference and a long-term followup demonstrates that macroprolactinemia develops before middle age and is likely a chronic condition. Polyethylene-glycol- (PEG-) precipitation method is widely used for screening macroprolactinemia, and gel filtration chromatography, protein A/G column, andI125-PRL binding studies are performed to confirm and clarify its nature. The cross-reactivity of macroprolactin varies widely according to the immunoassay systems. The epitope on PRL molecule recognized by the autoantibodies is located close to the binding site for PRL receptors, which may explain that macroprolactin has a lower biological activity. Hyperprolactinemia frequently seen in macroprolactinemic patients is due to the delayed clearance of autoantibody-bound PRL. When rats are immunized with rat pituitary PRL, anti-PRL autoantibodies are produced and hyperprolactinemia develops, mimicking macroprolactinemia in humans. Screening of macroprolactinemia is important for the differential diagnosis of hyperprolactinemia to avoid unnecessary examinations and treatments.

Download Full-text

A Carbon-Based Antifouling Nano-Biosensing Interface for Label-Free POCT of HbA1c

Biosensors ◽

10.3390/bios11040118 ◽

2021 ◽

Vol 11 (4) ◽

pp. 118

Author(s):

Zhenhua Li ◽

Jianyong Li ◽

Yanzhi Dou ◽

Lihua Wang ◽

Shiping Song

Keyword(s):

Electron Transfer ◽

Cyclic Voltammetry ◽

High Efficiency ◽

Dynamic Range ◽

Glycated Hemoglobin ◽

Electrochemical Impedance ◽

Label Free ◽

Electrochemical Biosensing ◽

Hemoglobin A ◽

Glycated Hemoglobin A

Electrochemical biosensing relies on electron transport on electrode surfaces. However, electrode inactivation and biofouling caused by a complex biological sample severely decrease the efficiency of electron transfer and the specificity of biosensing. Here, we designed a three-dimensional antifouling nano-biosensing interface to improve the efficiency of electron transfer by a layer of bovine serum albumin (BSA) and multi-walled carbon nanotubes (MWCNTs) cross-linked with glutaraldehyde (GA). The electrochemical properties of the BSA/MWCNTs/GA layer were investigated using both cyclic voltammetry and electrochemical impedance to demonstrate its high-efficiency antifouling nano-biosensing interface. The BSA/MWCNTs/GA layer kept 92% of the original signal in 1% BSA and 88% of that in unprocessed human serum after a 1-month exposure, respectively. Importantly, we functionalized the BSA/MWCNTs/GA layer with HbA1c antibody (anti-HbA1c) and 3-aminophenylboronic acid (APBA) for sensitive detection of glycated hemoglobin A (HbA1c). The label-free direct electrocatalytic oxidation of HbA1c was investigated by cyclic voltammetry (CV). The linear dynamic range of 2 to 15% of blood glycated hemoglobin A (HbA1c) in non-glycated hemoglobin (HbAo) was determined. The detection limit was 0.4%. This high degree of differentiation would facilitate a label-free POCT detection of HbA1c.

Download Full-text

Relationship of HbA1c Values to Retinopathy, Nephropathy, and Cardiovascular

International Journal of Drug Delivery Technology ◽

10.25258/ijddt.10.1.17 ◽

2020 ◽

Vol 10 (01) ◽

pp. 127-130

Author(s):

Aaya Hamid Al-Hakeem ◽

Hadeel Haider Saleh

Keyword(s):

Risk Factors ◽

Blood Pressure ◽

Diabetic Retinopathy ◽

Glycated Hemoglobin ◽

Microvascular Complications ◽

Diabetic Patients ◽

Glycaemia Control ◽

Relationship Of ◽

Hba1c Levels

A total of 50 patients aged 35-75 years From Al-Sader educational Hospital in Al-Najaf city was studied to determine the glycated hemoglobin risk factors with value creatinine and urea in serum and diabetic nephropathy. Diabetic patient were (35-45 years old) with HbA1c 7.9 % (60mmol/mol). Patient were (45–55 years old), glycated hemoglobin (HbA1c) andgt; 8.5 %. Patients between (55–65 years were glycated hemoglobin (HbA1c) andgt;10.5 %. HbA1c levels, lipid profile, level of Creatinine and urea in serum, family history, BMI, blood pressure, disease severity, and complications were determined. Most patients developed some grade of retinopathy (examined by an ophthalmologist) except those with HbA1c 6.7% (50mmol/mol). Diabetic patients aged (55–65 years old) with HbA1c 7.6% (60mmol/mol). Patients aged 56–75 years old of glycated hemoglobin (HbA1c) andgt;7% with poor glycaemia control ≥ 126mg/dL were assessed to classify diabetic retinopathy. HbA1c and GA are associated with nephropathy separately. Retinopathy and nephropathy may respond to different aspects of hyperglycemia. The GA found as a powerful indicator of microvascular complications same as HbA1c where long-term glycaemia is the risk factor.

Download Full-text

Label-free quantitation of glycated hemoglobin in single red blood cells by transient absorption microscopy and phasor analysis

Science Advances ◽

10.1126/sciadv.aav0561 ◽

2019 ◽

Vol 5 (5) ◽

pp. eaav0561 ◽

Cited By ~ 4

Author(s):

Pu-Ting Dong ◽

Haonan Lin ◽

Kai-Chih Huang ◽

Ji-Xin Cheng

Keyword(s):

Transient Absorption ◽

Glycated Hemoglobin ◽

Blood Glucose Concentration ◽

Label Free ◽

Absorption Imaging ◽

Fraction Distribution ◽

Boronate Affinity Chromatography ◽

Volume Blood ◽

Phasor Analysis

As a stable and accurate biomarker, glycated hemoglobin (HbA1c) is clinically used to diagnose diabetes with a threshold of 6.5% among total hemoglobin (Hb). Current methods such as boronate affinity chromatography involve complex processing of large-volume blood samples. Moreover, these methods cannot measure HbA1c fraction at single–red blood cell (RBC) level, thus unable to separate the contribution from other factors such as RBC lifetime. Here, we demonstrate a spectroscopic transient absorption imaging approach that is able to differentiate HbA1c from Hb on the basis of their distinct excited-state dynamics. HbA1c fraction inside a single RBC is derived quantitatively through phasor analysis. HbA1c fraction distribution of diabetic blood is apparently different from that of healthy blood. A mathematical model is developed to derive the long-term blood glucose concentration. Our technology provides a unique way to study heme modification and to derive clinically important information void of bloodstream glucose fluctuation.

Download Full-text

Quantitative microbiome profiling in lumenal and tissue samples with broad coverage and dynamic range via a single-step 16S rRNA gene DNA copy quantification and amplicon barcoding

10.1101/2020.01.22.914705 ◽

2020 ◽

Cited By ~ 3

Author(s):

Said R. Bogatyrev ◽

Rustem F. Ismagilov

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Microbial Diversity ◽

Dynamic Range ◽

Single Step ◽

Clinical Samples ◽

Marker Genes ◽

Rrna Gene ◽

Wide Dynamic Range ◽

Microbiome Profiling

ABSTRACTCurrent methods for detecting, accurately quantifying, and profiling complex microbial communities based on the microbial 16S rRNA marker genes are limited by a number of factors, including inconsistent extraction of microbial nucleic acids, amplification interference from contaminants and host DNA, different coverage of PCR primers utilized for quantification and sequencing, and potentially biases in PCR amplification rates among microbial taxa during amplicon barcoding. Here, we describe a single-step method that enables the quantification of microbial 16S rRNA gene DNA copies with wide dynamic range and broad microbial diversity, and simultaneous amplicon barcoding for quantitative 16S rRNA gene amplicon profiling of microbiota. The method is suitable for a variety of sample types and is robust in samples with low microbial abundance, including samples containing high levels of host mammalian DNA, as is common in human clinical samples. We demonstrate that our modification to the Earth Microbiome Project (EMP) V4 16S rRNA gene primers expands their microbial coverage while dramatically reducing non-specific mammalian mitochondrial DNA amplification, thus achieving wide dynamic range in microbial quantification and broad coverage for capturing high microbial diversity in samples with or without high host DNA background. The approach relies only on broadly available hardware (real-time PCR instruments) and standard reagents utilized for conventional 16S rRNA gene amplicon library preparation both of which make it amenable for immediate and widespread adoption. Simultaneous 16S rRNA gene DNA copy quantification and amplicon barcoding for multiplexed next-generation sequencing from the same analyzed sample, performed in a combined workflow, reduces the amount of sample needed and reduces time and reagent costs. Additionally, we demonstrate that using our modified 16S rRNA gene primers in a digital PCR (dPCR) format enables precise and exact microbial quantification in samples with very high host DNA background levels without the need for quantification standards. Potential future applications of this approach include: (1) quantitative microbiome profiling in human and animal microbiome research; (2) detection of monoinfections and profiling of polymicrobial infections in tissues, stool, and bodily fluids in human and veterinary medicine; (3) environmental sample analyses (e.g., soil and water); and (4) broad-coverage detection of microbial food contamination in products high in mammalian DNA, such as meat products. We predict that utilization of this approach primarily for quantitative microbiome profiling will be invaluable to microbiome studies, which have historically been limited to analysis of relative abundances of microbes.

Download Full-text

A Novel Label-Free Biosensor for Detection of HE4 in Urine Based on Localized Surface Plasmon Resonance and Protein G Directional Fixed

Journal of Nanomaterials ◽

10.1155/2020/8613240 ◽

2020 ◽

Vol 2020 ◽

pp. 1-7

Author(s):

Ruiqi Duan ◽

Mingrong Xi

Keyword(s):

Surface Plasmon Resonance ◽

Surface Plasmon ◽

Plasmon Resonance ◽

Localized Surface Plasmon Resonance ◽

Dynamic Range ◽

Localized Surface Plasmon ◽

Clinical Samples ◽

Protein G ◽

Label Free ◽

Early Screening

A non-invasive and more sensitive method for detection of HE4 is very important for the early screening and detection of ovarian carcinoma. In this study, we improved our previous localized surface plasmon resonance (LSPR) biosensor for detection of HE4 in urine to overcome disadvantages of conventional methods. Protein G directional fixed method was firstly used for LSPR biosensor to improved sensitivity, and standard HE4 and clinical samples were detected separately using this new biosensor. Compared to our previous LSPR biosensor, this new sensor was more sensitive, with other advantages as before. Under optimum conditions, this new biosensor could display a detection limit of 1 pM and wide dynamic range of 1 pM to 10,000 pM. This new biosensor was effective for detection of HE4 in urine of early ovarian cancer patients, without label and purification. To the best of our knowledge, this is first work to investigate LSPR biosensor for detection of tumor marker in urine, with great advantages and clinical application potentials.

Download Full-text

Waning antibody responses in COVID-19: what can we learn from the analysis of other coronaviruses?

Infection ◽

10.1007/s15010-021-01664-z ◽

2021 ◽

Author(s):

Ali Hamady ◽

JinJu Lee ◽

Zuzanna A. Loboda

Keyword(s):

Cross Reactivity ◽

Antibody Responses ◽

The Novel ◽

Incidence And Mortality ◽

Antibody Titres ◽

Human Coronaviruses ◽

Public Health Strategies ◽

Antibody Dynamics

Abstract Objectives The coronavirus disease 2019 (COVID-19), caused by the novel betacoronavirus severe acute respiratory syndrome 2 (SARS-CoV-2), was declared a pandemic in March 2020. Due to the continuing surge in incidence and mortality globally, determining whether protective, long-term immunity develops after initial infection or vaccination has become critical. Methods/Results In this narrative review, we evaluate the latest understanding of antibody-mediated immunity to SARS-CoV-2 and to other coronaviruses (SARS-CoV, Middle East respiratory syndrome coronavirus and the four endemic human coronaviruses) in order to predict the consequences of antibody waning on long-term immunity against SARS-CoV-2. We summarise their antibody dynamics, including the potential effects of cross-reactivity and antibody waning on vaccination and other public health strategies. At present, based on our comparison with other coronaviruses we estimate that natural antibody-mediated protection for SARS-CoV-2 is likely to last for 1–2 years and therefore, if vaccine-induced antibodies follow a similar course, booster doses may be required. However, other factors such as memory B- and T-cells and new viral strains will also affect the duration of both natural and vaccine-mediated immunity. Conclusion Overall, antibody titres required for protection are yet to be established and inaccuracies of serological methods may be affecting this. We expect that with standardisation of serological testing and studies with longer follow-up, the implications of antibody waning will become clearer.

Download Full-text

Critical angle reflection imaging for quantification of molecular interactions on glass surface

Nature Communications ◽

10.1038/s41467-021-23730-8 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Guangzhong Ma ◽

Runli Liang ◽

Zijian Wan ◽

Shaopeng Wang

Keyword(s):

Refractive Index ◽

Small Molecule ◽

Molecular Interactions ◽

Glass Surface ◽

Critical Angle ◽

Dynamic Range ◽

Refractive Index Change ◽

Label Free ◽

Index Change ◽

Sensing Range

AbstractQuantification of molecular interactions on a surface is typically achieved via label-free techniques such as surface plasmon resonance (SPR). The sensitivity of SPR originates from the characteristic that the SPR angle is sensitive to the surface refractive index change. Analogously, in another interfacial optical phenomenon, total internal reflection, the critical angle is also refractive index dependent. Therefore, surface refractive index change can also be quantified by measuring the reflectivity near the critical angle. Based on this concept, we develop a method called critical angle reflection (CAR) imaging to quantify molecular interactions on glass surface. CAR imaging can be performed on SPR imaging setups. Through a side-by-side comparison, we show that CAR is capable of most molecular interaction measurements that SPR performs, including proteins, nucleic acids and cell-based detections. In addition, we show that CAR can detect small molecule bindings and intracellular signals beyond SPR sensing range. CAR exhibits several distinct characteristics, including tunable sensitivity and dynamic range, deeper vertical sensing range, fluorescence compatibility, broader wavelength and polarization of light selection, and glass surface chemistry. We anticipate CAR can expand SPR′s capability in small molecule detection, whole cell-based detection, simultaneous fluorescence imaging, and broader conjugation chemistry.

Download Full-text