2. Academic Validation
  3. How to use Nile Red, a selective fluorescent stain for microalgal neutral lipids

How to use Nile Red, a selective fluorescent stain for microalgal neutral lipids

  • J Microbiol Methods. 2016 Sep;128:74-79. doi: 10.1016/j.mimet.2016.07.011.
Gibrán S Alemán-Nava 1,  Sara P Cuellar-Bermudez 2,  María Cuaresma 3,  Rouke Bosma 4,  Koenraad Muylaert 2,  Bruce E Ritmann 5,  Roberto Parra 6
Affiliations

Affiliations

  • 1 Escuela de Ingeniería y Ciencias, Tecnologico de Monterrey, Campus Monterrey, Ave. Eugenio Garza Sada 2501, Monterrey 64849, N.L., Mexico.
  • 2 Laboratory Aquatic Biology, KU Leuven Kulak, E. Sabbelaan 53, Kortrijk, Belgium.
  • 3 Algal Biotechnology Group, Centro de Investigación y Desarrollo de Recursos y Tecnologías Agroalimentarias (CIDERTA), Huelva University, Marine International Campus of Excellence (CEIMAR), Huelva, Spain.
  • 4 Wageningen University, Bioprocess Engineering, AlgaePARC, P.O. Box 16, 6700 AA, Wageningen, The Netherlands.
  • 5 Escuela de Ingeniería y Ciencias, Tecnologico de Monterrey, Campus Monterrey, Ave. Eugenio Garza Sada 2501, Monterrey 64849, N.L., Mexico; Swette Center for Environmental Biotechnology, Biodesign Institute, Arizona State University, PO Box 875701, Tempe, AZ 85287-5701, USA.
  • 6 Escuela de Ingeniería y Ciencias, Tecnologico de Monterrey, Campus Monterrey, Ave. Eugenio Garza Sada 2501, Monterrey 64849, N.L., Mexico. Electronic address: r.parra@itesm.mx.
PMID: 27432343 DOI: 10.1016/j.mimet.2016.07.011
Abstract

The use of Nile Red for rapid monitoring of the neutral lipid content in microalgae has gained interest over the last decade, since neutral lipids are feedstock for renewable transportation fuel. In this review, we discuss the main considerations needed to make an NR protocol reliable for staining neutral lipids in microalgae. Cell wall permeability must be enhanced by using stain carriers: DMSO (5% v/v to 25% v/v), glycerol (0.1 to 0.125mg/mL), or EDTA (3.0 to 3.8mg/mL). Temperatures between 30 and 40°C facilitate the diffusion of NR through the cell wall without incurring excess quenching. Good NR-lipid interaction requires using a low NR/cell ratio; the NR concentration must be between 0.25μg/mL and 2.0μg/mL, and the cell concentration >5×10(4)cells/mL. In order to have the maximum and stable NR fluorescence, it is necessary to scan the excitation/emission wavelengths for up to a 40-min of incubation time. We outline a five-step method to customize the Nile Red protocol to a specific strain: 1) Evaluate the strain's suitability by checking for the presence of neutral lipid, 2) Select of the best excitation/emission wavelength, 3) Optimization of incubation time, stain carrier, dye concentration, and temperature, 4) Prepare single-strain algal cultures with different lipid contents to calibrate NR fluorescence with neutral-lipid content, and 5) Correlate NR fluorescence intensity to neutral lipid content for the same strain. Once the protocol is customized, the NR method allows for rapid and reliable monitoring of neutral lipid content of a microalgae strain.

Keywords

Biodiesel; Microalgae; Neutral lipids; Nile Red; Protocol.

Figures
Products
嗨！很高兴为您提供帮助！

尊敬的 MCE 客户您好，
请您选择所在区域，我们将转接对应客服为您服务！

热门区域

A

B

C

F

G

H

J

L

N

Q

S

T

X

Y

Z

热 A B C F G H J L N Q S T X Y Z