44 research outputs found
Development of Nanostructured Triggerable Biosensors for Prostate Cancer Early Detection
No full textCancer remains a leading cause of global mortality, with Prostate Cancer (PCa) ranking as the second most common cancer in men. Early detection poses a significant challenge due to cancer's growth rate and limitations in current diagnostic assays. Despite Prostate Specific Antigen (PSA) being a longstanding biomarker for PCa diagnosis, its lack of specificity leads to a high rate of false positives. In response, alternative biomarkers like Prostate Specific Membrane Antigen (PSMA), detectable in urinary exosomes have emerged. Additionally, PSA in serum facilitates early detection of PCa recurrence post-prostatectomy. We developed two triggerable oligonucleotide-gold nanoparticle (AuNPs) architectures (AuNPs-Aggregates) that selectively disassembles in the presence of a protein biomarker (PSMA or PSA). The AuNPs-Aggregates were prepared by mixing two batches of AuNPs, functionalised with different ssDNA strands, including a sequence that contains an aptamer responsive to the protein marker. After in silico and in vitro sequences characterisation, I synthetized and functionalised AuNPs with thiolated-ssDNA strands. The PSMA responsive Aggregate is composed by 13 nm AuNPs, and is aimed to ex vivo early detection of PSMA in urinary exosomes. Its response was determined by monitoring changes in its size and optical properties. I obtained a significant specific PSMA response from the high femtomolar range, with purified PSMA protein, and from low picomolar range with the protein extract of PSMA positive exosomes. The second sensor, the AS2-US-AuNPs-Aggregate, is thought for an in vivo application to early detect PSA in the blood of totally resected patients and is composed by ultrasmall AuNPs, a size suitable for renal clearance. This Aggregate has a size of hundreds of nanometres, compatible with prolonged circulation time in bloodstream. In this way, the release of single AuNPs from the Aggregate upon aptamer recognition could permit its analysis in urine. The AS2-AuNPs-Aggregate response to PSA was monitored following its changes in size; its specific response started to be significant in the low fM range. Then, we compared the stability of AS2 annealed sequences to nucleases, both in their free dsDNA form and when integrated into the AS2-AuNPs-Aggregate. This nanostructure has the potential to be tailored for detecting various biomarkers, such as proteins or DNA, by modifying the nucleotide sequences involved in the recognition process. Furthermore, it offers flexibility in selecting the nanoparticles incorporated in the structure, with the possibility of different methods of detection
Libretto di sala - 2006 - Ensemble Festa Rustica
No full textGiorgio Matteoli, direttore e flauto dolceGianluca Belfiori Doro, contralt
Aptamer-based gold nanoparticle aggregates for ultrasensitive amplification-free detection of PSMA
No full textEarly diagnosis is one of the most important factors in determining the prognosis in cancer. Sensitive detection and quantification of tumour-specific biomarkers have the potential to improve significantly our diagnostic capability. Here, we introduce a triggerable aptamer-based nanostructure based on an oligonucleotide/gold nanoparticle architecture that selectively disassembles in the presence of the biomarker of interest; its optimization is based also on in-silico determination of the aptamer nucleotides interactions with the protein of interest. We demonstrate this scheme for the case of Prostate Specific Membrane Antigen (PSMA) and PSMA derived from PSMA-positive exosomes. We tested the disassembly of the system by diameter and count rate measurements in dynamic light scattering, and by inspection of its plasmon resonance shift, upon addition of PSMA, finding appreciable differences down to the sub-picomolar range; this points towards the possibility that this approach may lead to sensors competitive with diagnostic biochemical assays that require enzymatic amplification. More generally, this scheme has the potential to be applied to a broad range of pathologies with specific identified biomarkers
Complete Acid Ceramidase ablation prevents cancer-initiating cell formation in melanoma cells
No full textAcid ceramidase (AC) is a lysosomal cysteine hydrolase that catalyzes the conversion of ceramide into fatty acid and sphingosine. This reaction lowers intracellular ceramide levels and concomitantly generates sphingosine used for sphingosine-1-phosphate (S1P) production. Since increases in ceramide and consequent decreases of S1P reduce proliferation of various cancers, AC might offer a new target for anti-tumor therapy. Here we used CrispR-Cas9-mediated gene editing to delete the gene encoding for AC, ASAH1, in human A375 melanoma cells. ASAH1-null clones show significantly greater accumulation of long-chain saturated ceramides that are substrate for AC. As seen with administration of exogenous ceramide, AC ablation blocks cell cycle progression and accelerates senescence. Importantly, ASAH1-null cells also lose the ability to form cancer-initiating cells and to undergo self-renewal, which is suggestive of a key role for AC in maintaining malignancy and self-renewal of invasive melanoma cells. The results suggest that AC inhibitors might find therapeutic use as adjuvant therapy for advanced melanoma
Protezione da herpes genitale attraverso vaccinazione con glicoproteina B di herpes simplex di tipo 1.
No full textLa vaccinazione con la glicoproteina B del virus Herpes Simplex tipo 1 protegge contro l’infezione sessualmente trasmessa
No full textDelivery of an Immunogen with a FIV vector confers substantial protection against genital herpes.
No full textIn vivo delivery of herpes simplex virus 1 glycoprotein B to vaccinate against genital herpes
No full textLa vaccinazione con la glicoproteina B del virus Herpes Simplex tipo 1 protegge contro l’herpes genitale
No full textMeasuring pH in insulin secretory granules by phasor-based fluorescence lifetime imaging of a genetically encoded sensor
No full textIt is widely accepted that the pH of insulin granules is acidic, and that its active regulation during granule maturation plays a role in the process of insulin secretion by β-cells. Yet, a calibrated measurement of the absolute granule pH with organelle specificity is still lacking. To tackle this issue, we use the genetically encoded E1GFP pH reporter inserted into the C-peptide of proinsulin and expressed in Insulinoma 1E cells. Following verification of correct targeting of the E1GFP reporter in the insulin granules, phasor-based Fluorescence Lifetime Imaging Microscopy (FLIM) is applied to obtain a calibrated and probe-concentration-independent measurement of insulin-granule pH. Our results confirm the acidic nature of insulin granules under maintenance cell culture conditions, with an average luminal pH of ~5.8, and show that acidity is actively maintained, as evidenced by its near-neutralization upon treatment with the vacuolar H+-ATPase inhibitor concanamycin. Additionally, by ..
