scholarly journals Molecular Probe Optimization to Determine Cell Mortality in a Photosynthetic Organism (Microcystis aeruginosa) Using Flow Cytometry

10.3791/53036 ◽  
2016 ◽  
Author(s):  
Ian J. Chapman ◽  
Genoveva F. Esteban ◽  
Daniel J. Franklin
Keyword(s):  
Flow Cytometry ◽  
Microcystis Aeruginosa ◽  
Molecular Probe ◽  
Cell Mortality ◽  
Photosynthetic Organism ◽  
Probe Optimization

scholarly journals Effect of Zinc on Microcystis aeruginosa UTEX LB 2385 and Its Toxin Production

Toxins ◽  
2020 ◽  
Vol 12 (2) ◽  
pp. 92 ◽  
Author(s):  
Jose L. Perez ◽  
Tinchun Chu
Keyword(s):  
Flow Cytometry ◽  
Stress Response ◽  
Microcystis Aeruginosa ◽  
Stress Responses ◽  
Algal Blooms ◽  
Phase Contrast ◽  
Phase Contrast Microscopy ◽  
Zinc Stress ◽  
Cell Quota ◽  
Contrast Microscopy

Cyanobacteria harmful algal blooms (CHABs) are primarily caused by man-made eutrophication and increasing climate-change conditions. The presence of heavy metal runoff in affected water systems may result in CHABs alteration to their ecological interactions. Certain CHABs produce by-products, such as microcystin (MC) cyanotoxins, that have detrimentally affected humans through contact via recreation activities within implicated water bodies, directly drinking contaminated water, ingesting biomagnified cyanotoxins in seafood, and/or contact through miscellaneous water treatment. Metallothionein (MT) is a small, metal-sequestration cysteine rich protein often upregulated within the stress response mechanism. This study focused on zinc metal resistance and stress response in a toxigenic cyanobacterium, Microcystis aeruginosa UTEX LB 2385, by monitoring cells with (0, 0.1, 0.25, and 0.5 mg/L) ZnCl2 treatment. Flow cytometry and phase contrast microscopy were used to evaluate physiological responses in cultures. Molecular assays and an immunosorbent assay were used to characterize the expression of MT and MC under zinc stress. The results showed that the half maximal inhibitory concentration (IC50) was 0.25 mg/L ZnCl2. Flow cytometry and phase contrast microscopy showed morphological changes occurred in cultures exposed to 0.25 and 0.5 mg/L ZnCl2. Quantitative PCR (qPCR) analysis of selected cDNA samples showed significant upregulation of Mmt through all time points, significant upregulation of mcyC at a later time point. ELISA MC-LR analysis showed extracellular MC-LR (µg/L) and intracellular MC-LR (µg/cell) quota measurements persisted through 15 days, although 0.25 mg/L ZnCl2 treatment produced half the normal cell biomass and 0.5 mg/L treatment largely inhibited growth. The 0.25 and 0.5 mg/L ZnCl2 treated cells demonstrated a ~40% and 33% increase of extracellular MC-LR(µg/L) equivalents, respectively, as early as Day 5 compared to control cells. The 0.5 mg/L ZnCl2 treated cells showed higher total MC-LR (µg/cell) quota yield by Day 8 than both 0 mg/L ZnCl2 control cells and 0.1 mg/L ZnCl2 treated cells, indicating release of MCs upon cell lysis. This study showed this Microcystis aeruginosa strain is able to survive in 0.25 mg/L ZnCl2 concentration. Certain morphological zinc stress responses and the upregulation of mt and mcy genes, as well as periodical increased extracellular MC-LR concentration with ZnCl2 treatment were observed.

scholarly journals A Novel Quality Control Methodology on Screening out for Human Osteosarcoma Cell Adaptor Based on the Cell-SELEX Technique

2020 ◽  
Author(s):  
Hua Wang ◽  
Ji Liang ◽  
Yan Ma ◽  
Lei Zhou ◽  
Yufei Zhang ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Quality Control ◽  
Fluorescence Spectroscopy ◽  
Tumour Cell ◽  
Molecular Probe ◽  
Target Cells ◽  
Osteosarcoma Cell ◽  
Time Saving ◽  
Human Osteosarcoma ◽  
Human Osteosarcoma Cell

Abstract Background: We have successfully developed a novel molecular probe for recognition of human osteosarcoma cell using the cell-SELEX method. The study aims to establish an accurate, time-saving quality-monitoring method in screening for tumour cell adaptors in order to shorten the screening process and ensure the accurate preparation of the adaptor. Methods: Two kinds of human osteosarcoma cells (U2-OS, HOS) were selected as the forward screening target cells and human fibrosarcoma cells (HT-1080) as the reverse screening cells to screen the adaptors from the candidate oligonucleotide library. In each round of preparation of the library, PCR was optimised by using quantitative template concentration instead of percentage volume. Each round of forward screening was conducted with reverse screening; Fluorescence spectroscopy and flow cytometry were used to monitor and compare the aptamer libraries. Results: During quantitative PCR for U2-OS and HOS template, the results showed that the bands obtained from 14 cycles were bright and no non-specific amplification within the optimal template concentrations between 19.0 and 21.0ng/µl. Each round of forward screening was accompanied by reverse screening to accelerate the elimination of non-specific single-strand DNA (ssDNA). In the meanwhile, the adaptor groups were effectively purified specifically bounding to target cells. Besides, we observed that the fluorescence spectroscopy is more accurate, time-saving, and convenient for quality control compared with flow cytometry. Conclusion: The method proposed in the study is appropriate for the rapid screening out for human osteosarcoma cell adaptor. The quantitative template concentration, forward screening with back screening, and fluorescence spectroscopy are important methods for accurate preparation and quality control of tumour cell aptamers. It can provide scientific reference data for the amplification of dsDNAs in other sub-libraries.

Optimization of FDA–PI method using flow cytometry to measure metabolic activity of the cyanobacteria, Microcystis aeruginosa

2011 ◽  
Vol 36 (9-11) ◽  
pp. 424-429 ◽  
Author(s):  
Xi Xiao ◽  
Zhi-ying Han ◽  
Ying-xu Chen ◽  
Xin-qiang Liang ◽  
Hua Li ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Microcystis Aeruginosa ◽  
Metabolic Activity ◽  
Pi Method

scholarly journals Fluorescence Labeling of Circulating Tumor Cells with a Folate Receptor-Targeted Molecular Probe for Diffuse In Vivo Flow Cytometry

2020 ◽  
Vol 22 (5) ◽  
pp. 1280-1289 ◽  
Author(s):  
Roshani A. Patil ◽  
Madduri Srinivasarao ◽  
Mansoor M. Amiji ◽  
Philip S. Low ◽  
Mark Niedre
Keyword(s):  
Flow Cytometry ◽  
Tumor Cells ◽  
Circulating Tumor Cells ◽  
Folate Receptor ◽  
Molecular Probe ◽  
Fluorescence Labeling ◽  
In Vivo Flow Cytometry

Short term copper toxicity on Microcystis aeruginosa and Chlorella vulgaris using flow cytometry

2009 ◽  
Vol 94 (4) ◽  
pp. 255-264 ◽  
Author(s):  
Souad Hadjoudja ◽  
Chantal Vignoles ◽  
Véronique Deluchat ◽  
Jean-François Lenain ◽  
Anne-Hélène Le Jeune ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Microcystis Aeruginosa ◽  
Chlorella Vulgaris ◽  
Copper Toxicity ◽  
Short Term

scholarly journals Novel fluorescent antagonist as a molecular probe in A3 adenosine receptor binding assays using flow cytometry

2012 ◽  
Vol 83 (11) ◽  
pp. 1552-1561 ◽  
Author(s):  
Eszter Kozma ◽  
T. Santhosh Kumar ◽  
Stephanie Federico ◽  
Khai Phan ◽  
Ramachandran Balasubramanian ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Receptor Binding ◽  
Adenosine Receptor ◽  
Molecular Probe ◽  
Binding Assays ◽  
A3 Adenosine Receptor
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