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Micro-Droplet Detection Method for Measuring the Concentration of Alkaline Phosphatase-Labeled Nanoparticles in Fluorescence Microscopy
- Source :
- Sensors, Vol 17, Iss 11, p 2685 (2017), Sensors; Volume 17; Issue 11; Pages: 2685, Sensors (Basel, Switzerland)
- Publication Year :
- 2017
- Publisher :
- MDPI AG, 2017.
-
Abstract
- This paper developed and evaluated a quantitative image analysis method to measure the concentration of the nanoparticles on which alkaline phosphatase (AP) was immobilized. These AP-labeled nanoparticles are widely used as signal markers for tagging biomolecules at nanometer and sub-nanometer scales. The AP-labeled nanoparticle concentration measurement can then be directly used to quantitatively analyze the biomolecular concentration. Micro-droplets are mono-dispersed micro-reactors that can be used to encapsulate and detect AP-labeled nanoparticles. Micro-droplets include both empty micro-droplets and fluorescent micro-droplets, while fluorescent micro-droplets are generated from the fluorescence reaction between the APs adhering to a single nanoparticle and corresponding fluorogenic substrates within droplets. By detecting micro-droplets and calculating the proportion of fluorescent micro-droplets to the overall micro-droplets, we can calculate the AP-labeled nanoparticle concentration. The proposed micro-droplet detection method includes the following steps: (1) Gaussian filtering to remove the noise of overall fluorescent targets, (2) a contrast-limited, adaptive histogram equalization processing to enhance the contrast of weakly luminescent micro-droplets, (3) an red maximizing inter-class variance thresholding method (OTSU) to segment the enhanced image for getting the binary map of the overall micro-droplets, (4) a circular Hough transform (CHT) method to detect overall micro-droplets and (5) an intensity-mean-based thresholding segmentation method to extract the fluorescent micro-droplets. The experimental results of fluorescent micro-droplet images show that the average accuracy of our micro-droplet detection method is 0.9586; the average true positive rate is 0.9502; and the average false positive rate is 0.0073. The detection method can be successfully applied to measure AP-labeled nanoparticle concentration in fluorescence microscopy.
- Subjects :
- 0301 basic medicine
endocrine system
Materials science
Analytical chemistry
Nanoparticle
micro-droplet
02 engineering and technology
lcsh:Chemical technology
fluorescence microscopy
complex mixtures
Biochemistry
Article
alkaline phosphatase (AP)
Analytical Chemistry
Hough transform
law.invention
03 medical and health sciences
law
Fluorescence microscope
Humans
lcsh:TP1-1185
Electrical and Electronic Engineering
Instrumentation
Fluorescent Dyes
chemistry.chemical_classification
Biomolecule
spot detection
nanoparticles
technology, industry, and agriculture
Alkaline Phosphatase
021001 nanoscience & nanotechnology
Fluorescence
Thresholding
eye diseases
Atomic and Molecular Physics, and Optics
030104 developmental biology
Microscopy, Fluorescence
chemistry
Adaptive histogram equalization
0210 nano-technology
Biological system
Luminescence
Subjects
Details
- ISSN :
- 14248220
- Volume :
- 17
- Database :
- OpenAIRE
- Journal :
- Sensors
- Accession number :
- edsair.doi.dedup.....db619b4128e212c668bdc836d4a49b78