PMID- 33322812
OWN - NLM
STAT- PubMed-not-MEDLINE
LR  - 20201229
IS  - 2075-4418 (Print)
IS  - 2075-4418 (Electronic)
IS  - 2075-4418 (Linking)
VI  - 10
IP  - 12
DP  - 2020 Dec 12
TI  - A Light-Sheet-Based Imaaging Spectrometer to Characterize Acridine Orange 
      Fluorescence within Leukocytes.
LID - 10.3390/diagnostics10121082 [doi]
LID - 1082
AB  - Low-cost imaging systems that utilize exogenous fluorescent dyes, such as 
      acridine orange (AO), have recently been developed for use as point-of-care (POC) 
      blood analyzers. AO-based fluorescence imaging exploits variations in emission 
      wavelength within different cell types to enumerate and classify leukocyte 
      subpopulations from whole blood specimens. This approach to leukocyte 
      classification relies on accurate and reproducible colorimetric features, which 
      have previously been demonstrated to be highly dependent on the cell staining 
      protocols (such as specific AO concentration, timing, and pH). We have developed 
      a light-sheet-based fluorescence imaging spectrometer, featuring a spectral 
      resolution of 9 nm, with an automated spectral extraction algorithm as an 
      investigative tool to study the spectral features from AO-stained leukocytes. 
      Whole blood specimens were collected from human subjects, stained with AO using 
      POC methods, and leukocyte spectra were acquired on a cell-by-cell basis. The 
      post-processing method involves three steps: image segmentation to isolate 
      individual cells in each spectral image; image quality control to exclude cells 
      with low emission intensity, out-of-focus cells, and cellular debris; and the 
      extraction of spectra for each cell. An increase in AO concentration was 
      determined to contribute to the red-shift in AO-fluorescence, while varied pH 
      values did not cause a change in fluorescence. In relation to the spectra of 
      AO-stained leukocytes, there were corresponding red-shift trends associated with 
      dye accumulation within acidic vesicles and at increasing incubation periods. The 
      system presented here could guide future development of POC systems reliant on AO 
      fluorescence and colorimetric features to identify leukocyte subpopulations in 
      whole blood specimens.
FAU - Powless, Amy J
AU  - Powless AJ
AD  - Biomedical Engineering Department, University of Arkansas, Fayetteville, AR 
      72701, USA.
FAU - Prieto, Sandra P
AU  - Prieto SP
AD  - Biomedical Engineering Department, University of Arkansas, Fayetteville, AR 
      72701, USA.
FAU - Gramling, Madison R
AU  - Gramling MR
AD  - Pat Walker Health Center, University of Arkansas, Fayetteville, AR 72701, USA.
FAU - Chen, Jingyi
AU  - Chen J
AUID- ORCID: 0000-0003-0012-9640
AD  - Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville, 
      AR 72701, USA.
FAU - Muldoon, Timothy J
AU  - Muldoon TJ
AD  - Biomedical Engineering Department, University of Arkansas, Fayetteville, AR 
      72701, USA.
LA  - eng
GR  - N/A/Arkansas Biosciences Institute/
GR  - N/A/Arkansas Women's Giving Circle/
PT  - Journal Article
DEP - 20201212
PL  - Switzerland
TA  - Diagnostics (Basel)
JT  - Diagnostics (Basel, Switzerland)
JID - 101658402
PMC - PMC7763249
OTO - NOTNLM
OT  - acridine orange
OT  - fluorescence characteristics
OT  - imaging spectrometer
OT  - leukocytes
COIS- The authors declare no conflict of interest.
EDAT- 2020/12/17 06:00
MHDA- 2020/12/17 06:01
PMCR- 2020/12/12
CRDT- 2020/12/16 01:05
PHST- 2020/11/11 00:00 [received]
PHST- 2020/12/02 00:00 [revised]
PHST- 2020/12/08 00:00 [accepted]
PHST- 2020/12/16 01:05 [entrez]
PHST- 2020/12/17 06:00 [pubmed]
PHST- 2020/12/17 06:01 [medline]
PHST- 2020/12/12 00:00 [pmc-release]
AID - diagnostics10121082 [pii]
AID - diagnostics-10-01082 [pii]
AID - 10.3390/diagnostics10121082 [doi]
PST - epublish
SO  - Diagnostics (Basel). 2020 Dec 12;10(12):1082. doi: 10.3390/diagnostics10121082.