Nanosensors Flashcards
What is the size region of a nano sensor?
1nm to 100nm
What is a nano sensor?
Measures physical quantities (non electrical form) and converts to electrical form for discernible translation by humans
What are the current ways of detecting intracellular environments?
Free sensing dyes - toxic to cell, uneven cell loading, interference
Optodes (fibre optic with PVC and sensor on tip) - 100nm - creates holes in cell - apoptosis
What is PEBBLE?
Probe Encapsulated by Biologically Localised Embedding - flourophore surrounded by biologically inert capsule
What are the properties of PEBBLE?
20nm
97% cell viability
less than 1ms response time
What ions can PEBBLE detect?
K/Na/Ca/H/Mg/Cl - use Ionophore
How does PEBBLE detect glucose?
Contains enzyme (e.g. GoX) that converts glucose to H2O2 which can be measured
What is the inert capsule made of?
Acrylamide (+dextran) Sol Gel (+enzymes/PEG) Liquid Polymer (+additives/PEG)
What is a ratiometric O2 sensor?
Sol Gel matrix (silicon glass at low temp) Ruthenium Orange (sensor dye) - increase in O2 decreases sensitivity Oregon Green (reference) - no reaction to O2
Ratriometric sensing - ratio between sensor and reference intensity always stays constant - overcomes issues with uneven cell dye loading
How do we generate quantitive images using PEBBLE?
Superimpose differential phase contrast image with green and red confocal image (showing sensor and reference dyes)
How can we deliver the PEBBLE?
Gene Gun (holes - apoptosis) Macrophage phagocytosis (trapped in phagosome) Liposomal delivery (distributed randomly)
How do we produce PEBBLE devices?
Polymerase polymer (acrylamide) with flourophore
Boil off organic solvent used
Dry
Produces powder
How can we attach molecules to the outside?
Polymerase with monomers with free amine group - allows for attachment of molecules (e.g. biotin) using carboxyl, succimadyl group)
What is SelCA?
Sensing Living Cell Array - allows simultaneous detection of intracellular and extracellular ROS
Formation of MRC (Microtitre-plate Cell Retainers) with MREs (Micro Ring Elements with Gold (DTSSP - O2.) and Platinum (NO selective membrane) allows for extracellular sensing
What is CFS?
Myalgic Encephalopathy - chronic fatigue, muscle pain
Reduced bioenergic function (ability of cells to produce sufficient energy) shown with fMRI (shows brain activity via blood flow)
Linked to viral infection/accident/operations
Hypothesis that increased PD Kinase in CFS patients, breaks down PDC (involved in aerobic respiration, converting pyruvate to acetyl CoA). As a result increased anaerobic respiration (produces lactic acid - muscle fatigue)
What is DCA?
Dicholoracetate - inhibits PD K - used to treat lactic acidosis normally.
What is a pH sensing PEBBLE?
FITC - pH sensitive (increase in pH increases fluorescence)
AlexaFlour568 - reference
What studies have been done using the pH PEBBLE on CFS patients?
pH PEBBLE inserted (liposomal delivery) into myocytes from patients with CFS - decrease in fluorescence due to decrease in pH
Tested in cells soaked with DCA - increase in fluorescence due to increase in pH
What is the issue with DCA?
Toxic over time
