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Bright near-infrared fluorescence bio-labeling with a biliprotein triad
摘要： Biliproteins have extended the spectral range of fluorescent proteins into the near-infrared region (NIR, 700–770 nm) of maximal transmission of most tissues and are also favorable for multiplex labeling. Their application, however, presents considerable challenges to increase their stability under physiological conditions and, in particular, to increase their brightness while maintaining the emission in near-infrared regions: their fluorescence yield generally decreases with increasing wavelengths, and their effective brightness depends strongly on the environmental conditions. We report a fluorescent biliprotein triad, termed BDFP1.1:3.1:1.1, that combines a large red-shift (722 nm) with high brightness in mammalian cells and high stability under changing environmental conditions. It is fused from derivatives of the phycobilisome core subunits, ApcE2 and ApcF2. These two subunits are induced by far-red light (FR, 650–700 nm) in FR acclimated cyanobacteria. Two BDFP1.1 domains engineered from ApcF2 covalently bind biliverdin that is accessible in most cells. The soluble BDFP3 domain, engineered from ApcE2, binds phytochromobilin non-covalently, generating BDFP3.1. This phytochromobilin chromophore was added externally; it is readily generated by an improved synthesis in E. coli and subsequent extraction. Excitation energy absorbed in the FR by covalently bound biliverdins in the two BDFP1.1 domains is transferred via fluorescence resonance energy transfer (FRET) to the non-covalently bound phytochromobilin in the BDFP3.1 domain fluorescing in the NIR around 720 nm. Labeling of a variety of proteins by fusion to the biliprotein triad is demonstrated in prokaryotic and mammalian cells, including human cell lines.
关键词： Bioimaging,Biliprotein,FRET,Allophycocyanin,Biomarker,Biliverdin
更新于2025-09-10 09:29:36
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Diagnostic Biosensors in Medicine- a Review
摘要： Biosensors consist of a biocatalyst that can detect a biological element and a transducer which can convert the combination event of the biocatalyst and the biological element into a detectable parameter. The biocatalyst can be biomolecules like, enzyme, DNA, RNA, metabolites, cells, oligonucleotides etc., and the transducers can be electrochemical, optical, piezoelectric, acoustics, calorimetric etc. In disease diagnostics biosensors utilizing immobilized cells, enzymes, nucleic acids have come into the field in recent years. Nanobiosensors exploiting the ultrasmall size and unique properties have also been used for engineering disease diagnostic biosensors. The use of biosensors can rapidly assess the health status, onset of the disease and its progression and can help to plan treatment for many diseases with the aid of multidisciplinary combination of chemistry, medical science and nanotechnology. The devices are cost effective, highly sensitive, rapid, user friendly and can be produced in bulk for human use. This review focuses on the different biosensors for the diagnosis of three major diseases like diabetes, cardiovascular disease and cancer.
关键词： nanobiosensors,cancer,diabetes,cardiovascular disease,disease biomarker detection,Biosensors
更新于2025-09-10 09:29:36
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[Methods in Molecular Biology] FOXO Transcription Factors Volume 1890 (Methods and Protocols) || Immunofluorescence Analysis by Confocal Microscopy for Detecting Endogenous FOXO
摘要： Cancer cells are known to inactivate tumor suppressor proteins by triggering their anomalous subcellular location. It has been well established that the aberrant location of FOXO proteins is linked to tumor formation, progression of the same, or resistance to anti-neoplastic treatment. Furthermore, the abnormal location of FOXO has also been considered a potential biomarker for diabetic complications or longevity in different organisms. Here, we describe the immunodetection of endogenous FOXO by confocal microscopy, which can be used as a chemical tool to quantify FOXO expression levels, its cellular location, and even its active/inactive forms with relevant antibodies.
关键词： Immunodetection,Nuclear translocation,FOXO proteins,Confocal microscopy,Biomarker
更新于2025-09-10 09:29:36
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Multiplexed determination of intracellular messenger RNA by using a graphene oxide nanoprobe modified with target-recognizing fluorescent oligonucleotides
摘要： A multiplexed graphene oxide (GO) fluorescent nanoprobe is described for quantification and imaging of messenger RNAs (mRNAs) in living cells. The recognizing oligonucleotides (with sequences complementary to those of target the fluorescence of the recognizing mRNAs) were labeled with different fluorescent dyes. If adsorbed on GO, oligonucleotides is quenched. After having penetrated living cells, the oligonucleotides bind to target mRNAs and dissociate from GO. This leads to the recovery of fluorescence. Using different fluorescent dyes, various intracellular mRNAs can be simultaneously imaged and quantified by a high content analysis within a short period of time. Actin mRNA acts as the internal control. This GO-based nanoprobe allows mRNA mimics to be determined within an analytical range from 1 to 400 nM and a detection limit as low as 0.26 nM. Up to 3 intracellular mRNAs (C-myc, TK1, and actin) can be detected simultaneously in a single living cell. Hence, this nanoprobe enables specific distinction of intracellular mRNA expression levels in cancerous and normal cells. It can be potentially applied as a tool for detection of cancer progression and diagnosis.
关键词： Fluorescence resonance energy transfer (FRET),Cancer biomarker,Actin mRNA,Fluorometric detection,High content analysis,Cancer diagnosis
更新于2025-09-10 09:29:36
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Liquid Crystals: A Novel Approach for Cancer Detection and Treatment
摘要： Liquid crystals are defined as the fourth state of matter forming between solid and liquid states. Earlier the applications of liquid crystals were confined to electronic instruments, but recent research findings suggest multiple applications of liquid crystals in biology and medicine. Here, the purpose of this review article is to discuss the potential biological impacts of liquid crystals in the diagnosis and prognosis of cancer along with the risk assessment. In this review, we also discussed the recent advances of liquid crystals in cancer biomarker detection and treatment in multiple cell line models. Cases reviewed here will demonstrate that cancer diagnostics based on the multidisciplinary technology and intriguingly utilization of liquid crystals may become an alternative to regular cancer detection methodologies. Additionally, we discussed the formidable challenges and problems in applying liquid crystal technologies. Solving these problems will require great effort and the way forward is through the multidisciplinary collaboration of physicists, biologists, chemists, material-scientists, clinicians, and engineers. The triumphant outcome of these liquid crystals and their applications in cancer research would be convenient testing for the detection of cancer and may result in treating the cancer patients non-invasively.
关键词： biosensor,biomarker,antitumor drug,liquid crystals,cancers
更新于2025-09-10 09:29:36
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P2.06-28 Assessment of Chest Wall Motion Using Structured Light Plethysmography (SLP) in Mesothelioma and Benign Pleural Disease
摘要： Our study population presented a median age of 60 years (53 -71 yrs), most were male 64.8 %, with asbestos exposure in 57.2% of cases and epithelioid histology in 85.7%. Almost half of the patients received gemcitabine as ?rst-line treatment (n?43; 47.3%). Overall PFS was 14.1 months (95% CI 9.8e18.4 months,) while PFS in patients receiving gemcitabine treatment was 6.8 months (95% CI 5.4e8.2 months). Several factors were associated to an improved PFS, including female vs. male sex (7.2 vs 6.6 months, p?0.062) and epithelial vs. sarcomatoid histology (7.2 vs 3.8 months, p?0.035) after gemcitabine treatment in univariate analysis. In the multivariate analysis, sex (HR 3.06, p?0.085) and no wood-smoke exposure (HR 2.55, p?0.092) were independently associated to gemcitabine treatment. Patients with high expression levels of RRM1 showed better PFS compared to patients with low expression levels (7.6 vs 5.5 months, p?0.049) in univariate analysis; with a HR of 0.204 (0.050e0.831) p?0.026 in multivariate analysis. Conversely, the M2 subunit (RRM2) did not show any significant associations (7.6 vs 6.6 months, p?0.363). Conclusion: Our results suggest that high protein expression levels of RRM1 could potentially serve as a biomarker of response to gemcitabine treatment in patients with advanced stage MPM.
关键词： response biomarker,RRM2,RRM1
更新于2025-09-09 09:28:46
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Targeted Feature Extraction in MALDI Mass Spectrometry Imaging to Discriminate Proteomic Profiles of Breast and Ovarian Cancer
摘要： Purpose: To develop a mass spectrometry imaging (MSI) based workflow for extracting m/z values related to putative protein biomarkers and using these for reliable tumor classification. Experimental design: Given a list of putative breast and ovarian cancer biomarker proteins, we extracted a set of related m/z values from heterogeneous MSI datasets derived from formalin-fixed paraffin-embedded tissue material. Based on these features, a linear discriminant analysis classification model was trained to discriminate the two tumor types. Results: We show that the discriminative power of classification models based on the extracted features is increased compared to the automatic training approach, especially when classifiers are applied to spectral data acquired under different conditions (instrument, preparation, laboratory). Conclusions and clinical relevance: We obtained robust classification models not confounded by technical variation between MSI measurements. This supports the assumption that the classification of the respective tumor types is based on biological rather than technical differences, and that the selected features are related to the proteomic profiles of the tumor types under consideration.
关键词： feature extraction,tumor typing,MALDI MSI,tissue classification,biomarker proteins
更新于2025-09-09 09:28:46
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Hyperreflective Intraretinal Foci as an OCT Biomarker of Retinal Inflammation in Diabetic Macular Edema
摘要： We read with great interest the article titled ‘Association Between Soluble CD14 in the Aqueous Humor and Hyperre?ective Foci on Optical Coherence Tomography in Patients with Diabetic Macular Edema’ by Lee et al. The authors investigated the expression of soluble CD14 (sCD14) in aqueous humor (AH) samples of patients with diabetic macular edema (DME), and the presence of hyperre?ective foci (HF) on spectral-domain optical coherence tomography (SD-OCT) in these eyes. The authors found both higher levels of sCD14, and increased number of HF in the inner retina compared to controls, and concluded that, since sCD14 is released by retinal microglia, HF might represent aggregates of activated microglial cells in DME eyes. The presence of and changes in OCT HF, also called hyperre?ective spots, have been previously described in detail in diabetic eyes and reported to depend on activated microglia. DME-affected eyes treated with intravitreal anti-VEGF and steroids have also been investigated, demonstrating a reduction of HF number with both treatments—con?rming HF as a retinal in?ammatory biomarker. To improve this observation, we also investigated diabetic eyes in a large population to identify HF-speci?c characteristics, a topic not suf?ciently addressed by Lee et al. In particular, we disagree with the statement that HF, as aggregates of retinal microglial cells, have the same re?ectivity as the retinal pigment epithelium (RPE), whereas it is more similar to that of the retinal nerve ?ber layer (RNFL). Other clinical characteristics (size < 30 lm, absence of back-shadowing, presence in both inner and outer retinal layers in DME) allow differentiation of HF, as an in?ammatory biomarker, from hard exudates, small intraretinal hemorrhages, microaneurysms, and tiny capillaries, which also appear as OCT hyperre?ective foci (spots). The correct identi?cation of in?ammation-driven HF from other forms of hyperre?ective material is relevant when using this new biomarker in the follow-up of any DME treatment. As consequence, any investigation of HF on OCT in diabetic eyes should be more precise to avoid inconsistent results.
关键词： Hyperre?ective intraretinal foci,Retinal in?ammation,Diabetic macular edema,OCT biomarker
更新于2025-09-09 09:28:46
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Aluminum(III) triggered aggregation-induced emission of glutathione-capped copper nanoclusters as a fluorescent probe for creatinine
摘要： Glutathione-capped copper nanoclusters (CuNCs) are presented that display aggregation-induced emission (AIE). This feature was exploited for selective and sensitive quantification of creatinine (CRN) which is an important diagnostic parameter. In the presence of Al3+ ions, such CuNCs rapidly aggregate, and this induces enhanced a red emission. The AIE nature of CuNCs was proven via TEM and fluorimetry. On addition of CRN, the coordination between CRN and Al3+ ions led to the quenching of fluorescence due to weakening the AIE. The best fluorescence intensity was measured at excitation/emission peaks of 360/585 nm. Quenched fluorescence intensity showed a linear dependence on the concentrations of CRN in the range of 2.5–34 μgL?1 with a detection limit of 0.63 μgL?1. The sensing mechanism of probe for CRN detection is discussed. The probe was applied to the determination of CRN in spiked human serum samples and gave satisfactory results.
关键词： Nanosensor,Renal biomarker,Serum analysis,Fluorescent nanomaterials,Metal nanoclusters,Nanoprobe,Paired t-test,Fluorometry,Real sample analysis
更新于2025-09-04 15:30:14
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SERS-Based Quantification of PSMA in Tissue Microarrays Allows Effective Stratification of Patients with Prostate Cancer
摘要： Prostate specific membrane antigen (PSMA), a type II membrane protein, is an attractive biomarker that has been validated clinically for the diagnosis of prostate cancer. In this study, we developed surface-enhanced Raman scattering (SERS) nanoprobes for PSMA detection and quantification at the single-cell level on prostate cancer cells. The cells were targeted employing SERS nanoprobes that consisted of gold nanostars functionalized with PSMA aptamer molecules. We were able to quantify picomolar concentrations of soluble PSMA protein and used the resulting calibration curve to estimate the expression of PSMA on the surface of the prostate cancer cell, LNCaP, at the single-cell level. Importantly, we employed these SERS tags to stratify prostate cancer patients by assessing PSMA expression in tissues contained in a prostate tissue microarray. The stratification results clearly correlated PSMA expression to recommended therapy groups, rendering the described method as an effective tool to aid in designing personalized therapeutic protocols. Benchmarking detection sensitivity against immunofluorescence staining and comparing stratification results obtained with the two methods allowed us to validate our novel approach against standard practices. On the basis of these results, we confirm the validity of PSMA as an effective biomarker for prostate cancer patient evaluation and propose SERS-based diagnostic techniques as integrative methods for the assessment of disease stage and the identification of effective therapeutic protocols.
关键词： aptamer,tissue microarray,surface-enhanced Raman scattering,PSMA,Prostate specific membrane antigen,SERS,nanoprobes,prostate cancer,biomarker,gold nanostars
更新于2025-09-04 15:30:14