Skip to main content
SpringerLink
Log in

Application of NBD-Labeled Lipids in Membrane and Cell Biology

  • Chapter
  • First Online:
Fluorescent Methods to Study Biological Membranes

Part of the book series: Springer Series on Fluorescence ((SS FLUOR,volume 13))

Abstract

The fluorescent NBD group has come a long way in terms of biological applications since its discovery a few decades back. Although the field of fluorescently labeled lipids has grown over the years with the introduction of new fluorescent labels, NBD-labeled lipids continue to be a popular choice in membrane and cell biological studies due to desirable fluorescence characteristics of the NBD group. In this chapter, we discuss the application of NBD-labeled lipids in membrane and cell biology taking representative examples with specific focus on the biophysical basis underlying such applications.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Chapter
USD 29.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 259.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 329.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 329.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Abbreviations

6-NBD-PC:

1-Palmitoyl-2-(6-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]hexanoyl)-sn-glycero-3-phosphocholine

12-NBD-PC:

1-Palmitoyl-2-(12-[N-(7-nitrobenz-2-oxa-1,3-diazol-yl)amino]dodecanoyl)-sn-glycero-3-phosphocholine

6-NBD-CM:

6-([N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]hexanoyl)sphingosine

6-NBD-SM:

6-([N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]hexanoyl) sphingosylphosphocholine

25-NBD-cholesterol:

25-[N-[(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-methyl]amino]-27-norcholesterol

DOPC:

Dioleoyl-sn-glycero-3-phosphocholine

DPPC:

1,2-Dipalmitoyl-sn-glycero-3-phosphocholine

FRAP:

Fluorescence recovery after photobleaching

NBD:

7-Nitrobenz-2-oxa-1,3-diazol-4-yl

NBD-PE:

N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine

NBD-PS:

1,2-Dioleoyl-sn-glycero-3-phospho-l-serine-N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)

POPC:

1-Palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine

REES:

Red edge excitation shift

References

  1. Simons K, Toomre D (2000) Lipid rafts and signal transduction. Nat Rev Mol Cell Biol 1:31–39

    Article  CAS  Google Scholar 

  2. Pucadyil TJ, Chattopadhyay A (2007) Cholesterol: a potential therapeutic target in Leishmania infection? Trends Parasitol 23:49–53

    Article  CAS  Google Scholar 

  3. Riethmüller J, Riehle A, Grassmé H, Gulbins E (2006) Membrane rafts in host-pathogen interactions. Biochim Biophys Acta 1758:2139–2147

    Article  Google Scholar 

  4. Zimmerberg J (2006) Membrane biophysics. Curr Biol 16:R272–R276

    Article  CAS  Google Scholar 

  5. van Meer G, de Kroon AIPM (2011) Lipid map of the mammalian cell. J Cell Sci 124:5–8

    Article  Google Scholar 

  6. Chattopadhyay A (ed.) (2002) Lipid probes in membrane biology. Chem Phys Lipids 116:1–188

    Google Scholar 

  7. Eggeling C, Ringemann C, Medda R, Schwarzmann G, Sandhoff K, Polyakova S, Belov VN, Hein B, von Middendorff C, Schönle A, Hell SW (2009) Direct observation of the nanoscale dynamics of membrane lipids in a living cell. Nature 457:1159–1163

    Article  CAS  Google Scholar 

  8. Chattopadhyay A (1990) Chemistry and biology of N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-labeled lipids: fluorescent probes of biological and model membranes. Chem Phys Lipids 53:1–15

    Article  CAS  Google Scholar 

  9. Chattopadhyay A, Mukherjee S, Raghuraman H (2002) Reverse micellar organization and dynamics: a wavelength-selective fluorescence approach. J Phys Chem B 106:13002–13009

    Article  CAS  Google Scholar 

  10. Chattopadhyay A, London E (1988) Spectroscopic and ionization properties of N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-labeled lipids in model membranes. Biochim Biophys Acta 938:24–34

    Article  CAS  Google Scholar 

  11. Fery-Forgues S, Fayet JP, Lopez A (1993) Drastic changes in the fluorescence properties of NBD probes with the polarity of the medium: involvement of a TICT state? J Photochem Photobiol A 70:229–243

    Article  CAS  Google Scholar 

  12. Lin S, Struve WS (1991) Time-resolved fluorescence of nitrobenzoxadiazole-aminohexanoic acid: effect of intermolecular hydrogen-bonding on non-radiative decay. Photochem Photobiol 54:361–365

    Article  CAS  Google Scholar 

  13. Mukherjee S, Chattopadhyay A, Samanta A, Soujanya T (1994) Dipole moment change of NBD group upon excitation studied using solvatochromic and quantum chemical approaches: implications in membrane research. J Phys Chem 98:2809–2812

    Article  CAS  Google Scholar 

  14. Rawat SS, Chattopadhyay A (1999) Structural transition in the micellar assembly: a fluorescence study. J Fluoresc 9:233–244

    Article  CAS  Google Scholar 

  15. Koval M, Pagano RE (1990) Sorting of an internalized plasma membrane lipid between recycling and degradative pathways in normal and Niemann-Pick, type A fibroblasts. J Cell Biol 111:429–442

    Article  CAS  Google Scholar 

  16. Pagano RE, Sleight RG (1985) Defining lipid transport pathways in animal cells. Science 229:1051–1057

    Article  CAS  Google Scholar 

  17. Sparrow CP, Patel S, Baffic J, Chao Y-S, Hernandez M, Lam M-H, Montenegro J, Wright SD, Detmers PA (1999) A fluorescent cholesterol analog traces cholesterol absorption in hamsters and is esterified in vivo and in vitro. J Lipid Res 40:1747–1757

    CAS  Google Scholar 

  18. van Meer G, Stelzer EHK, Wijnaendts-van-Resandt RW, Simons K (1987) Sorting of sphingolipids in epithelial (Madin-Darby canine kidney) cells. J Cell Biol 105:1623–1635

    Article  Google Scholar 

  19. Mukherjee S, Zha X, Tabas I, Maxfield FR (1998) Cholesterol distribution in living cells: fluorescence imaging using dehydroergosterol as a fluorescent cholesterol analog. Biophys J 75:1915–1925

    Article  CAS  Google Scholar 

  20. Scheidt HA, Müller P, Herrmann A, Huster D (2003) The potential of fluorescent and spin-labeled steroid analogs to mimic natural cholesterol. J Biol Chem 278:45563–45569

    Article  CAS  Google Scholar 

  21. Bhattacharyya K, Bagchi B (2000) Slow dynamics of constrained water in complex geometries. J Phys Chem A 104:10603–10613

    Article  CAS  Google Scholar 

  22. Chattopadhyay A (2003) Exploring membrane organization and dynamics by the wavelength-selective fluorescence approach. Chem Phys Lipids 122:3–17

    Article  CAS  Google Scholar 

  23. Demchenko AP (2008) Site-selective red-edge effects. Methods Enzymol 450:59–78

    Article  CAS  Google Scholar 

  24. Haldar S, Chaudhuri A, Chattopadhyay A (2011) Organization and dynamics of membrane probes and proteins utilizing the red edge excitation shift. J Phys Chem B 115:5693–5706

    Article  CAS  Google Scholar 

  25. Mukherjee S, Chattopadhyay A (1995) Wavelength-selective fluorescence as a novel tool to study organization and dynamics in complex biological systems. J Fluoresc 5:237–246

    Article  CAS  Google Scholar 

  26. Haldar S, Chattopadhyay A (2007) Dipolar relaxation within the protein matrix of the green fluorescent protein: a red edge excitation shift study. J Phys Chem B 111:14436–14439

    Article  CAS  Google Scholar 

  27. Stubbs CD, Ho C, Slater SJ (1995) Fluorescence techniques for probing water penetration into lipid bilayers. J Fluoresc 5:19–28

    Article  CAS  Google Scholar 

  28. Chattopadhyay A, Mukherjee S (1993) Fluorophore environments in membrane-bound probes: a red edge excitation shift study. Biochemistry 32:3804–3811

    Article  CAS  Google Scholar 

  29. Abrams FS, London E (1993) Extension of the parallax analysis of membrane penetration depth to the polar region of model membranes: use of fluorescence quenching by a spin-label attached to the phospholipid polar headgroup. Biochemistry 32:10826–10831

    Article  CAS  Google Scholar 

  30. Chattopadhyay A, London E (1987) Parallax method for direct measurement of membrane penetration depth utilizing fluorescence quenching by spin-labeled phospholipids. Biochemistry 26:39–45

    Article  CAS  Google Scholar 

  31. Mitra B, Hammes GG (1990) Membrane-protein structural mapping of chloroplast coupling factor in asolectin vesicles. Biochemistry 29:9879–9884

    Article  CAS  Google Scholar 

  32. Mukherjee S, Raghuraman H, Dasgupta S, Chattopadhyay A (2004) Organization and dynamics of N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-labeled lipids: a fluorescence approach. Chem Phys Lipids 127:91–101

    Article  CAS  Google Scholar 

  33. Wolf DE, Winiski AP, Ting AE, Bocian KM, Pagano RE (1992) Determination of the transbilayer distribution of fluorescent lipid analogues by nonradiative fluorescence energy transfer. Biochemistry 31:2865–2873

    Article  CAS  Google Scholar 

  34. Chattopadhyay A, Mukherjee S (1999) Red edge excitation shift of a deeply embedded membrane probe: implications in water penetration in the bilayer. J Phys Chem B 103:8180–8185

    Article  CAS  Google Scholar 

  35. Rawat SS, Mukherjee S, Chattopadhyay A (1997) Micellar organization and dynamics: a wavelength-selective fluorescence approach. J Phys Chem B 101:1922–1929

    Article  CAS  Google Scholar 

  36. Raghuraman H, Chattopadhyay A (2007) Orientation and dynamics of melittin in membranes of varying composition utilizing NBD fluorescence. Biophys J 92:1271–1283

    Article  CAS  Google Scholar 

  37. Chattopadhyay A, Mukherjee S (1999) Depth-dependent solvent relaxation in membranes: wavelength-selective fluorescence as a membrane dipstick. Langmuir 15:2142–2148

    Article  CAS  Google Scholar 

  38. Huster D, Müller P, Arnold K, Herrmann A (2001) Dynamics of membrane penetration of the fluorescent 7-nitrobenz-2-oxa-1,3-diazol-4-yl (NBD) group attached to an acyl chain of phosphatidylcholine. Biophys J 80:822–831

    Article  CAS  Google Scholar 

  39. Huster D, Müller P, Arnold K, Herrmann A (2003) Dynamics of lipid chain attached fluorophore 7-nitrobenz-2-oxa-1,3-diazol-4-yl (NBD) in negatively charged membranes determined by NMR spectroscopy. Eur Biophys J 32:47–54

    CAS  Google Scholar 

  40. Loura LMS, Ramalho JPP (2007) Location and dynamics of acyl chain NBD-labeled phosphatidylcholine (NBD-PC) in DPPC bilayers. A molecular dynamics and time-resolved fluorescence anisotropy study. Biochim Biophys Acta 1768:467–478

    Article  CAS  Google Scholar 

  41. Raghuraman H, Shrivastava S, Chattopadhyay A (2007) Monitoring the looping up of acyl chain labeled NBD lipids in membranes as a function of membrane phase state. Biochim Biophys Acta 1768:1258–1267

    Article  CAS  Google Scholar 

  42. Tsukanova V, Grainger DW, Salesse C (2002) Monolayer behavior of NBD-labeled phospholipids at the air/water interface. Langmuir 18:5539–5550

    Article  CAS  Google Scholar 

  43. Mukherjee S, Soe TT, Maxfield FR (1999) Endocytic sorting of lipid analogues differing solely in the chemistry of their hydrophobic tails. J Cell Biol 144:1271–1284

    Article  CAS  Google Scholar 

  44. Fernandes F, Loura LMS, Koehorst R, Spruijt RB, Hemminga MA, Fedorov A, Prieto M (2004) Quantification of protein-lipid selectivity using FRET: application to the M13 major coat protein. Biophys J 87:344–352

    Article  CAS  Google Scholar 

  45. Mazères S, Schram V, Tocanne J-F, Lopez A (1996) 7-Nitrobenz-2-oxa-1,3-diazole-4-yl-labeled phospholipids in lipid membranes: differences in fluorescence behavior. Biophys J 71:327–335

    Article  Google Scholar 

  46. Crowley KS, Reinhart GD, Johnson AE (1993) The signal sequence moves through a ribosomal tunnel into a noncytoplasmic aqueous environment at the ER membrane early in translocation. Cell 73:1101–1115

    Article  CAS  Google Scholar 

  47. Shatursky O, Heuck AP, Shepard LA, Rossjohn J, Parker MW, Johnson AE, Tweten RK (1999) The mechanism of membrane insertion for a cholesterol-dependent cytolysin: a novel paradigm for pore-forming toxins. Cell 99:293–299

    Article  CAS  Google Scholar 

  48. Chapman CF, Liu Y, Sonek GJ, Tromberg BJ (1995) The use of exogenous fluorescent probes for temperature measurements in single living cells. Photochem Photobiol 62:416–425

    Article  CAS  Google Scholar 

  49. McIntyre JC, Sleight RG (1991) Fluorescence assay for phospholipid membrane asymmetry. Biochemistry 30:11819–11827

    Article  CAS  Google Scholar 

  50. Chaudhuri A, Chattopadhyay A (2011) Transbilayer organization of membrane cholesterol at low concentrations: implications in health and disease. Biochim Biophys Acta 1808:19–25

    Article  CAS  Google Scholar 

  51. Mukherjee S, Chattopadhyay A (1996) Membrane organization at low cholesterol concentrations: a study using 7-nitrobenz-2-oxa-1,3-diazol-4-yl-labeled cholesterol. Biochemistry 35:1311–1322

    Article  CAS  Google Scholar 

  52. Mukherjee S, Chattopadhyay A (2005) Monitoring cholesterol organization in membranes at low concentrations utilizing the wavelength-selective fluorescence approach. Chem Phys Lipids 134:79–84

    Article  CAS  Google Scholar 

  53. Rukmini R, Rawat SS, Biswas SC, Chattopadhyay A (2001) Cholesterol organization in membranes at low concentrations: effects of curvature stress and membrane thickness. Biophys J 81:2122–2134

    Article  CAS  Google Scholar 

  54. Pucadyil TJ, Mukherjee S, Chattopadhyay A (2007) Organization and dynamics of NBD-labeled lipids in membranes analyzed by fluorescence recovery after photobleaching. J Phys Chem B 111:1975–1983

    Article  CAS  Google Scholar 

  55. Loura LMS, Prieto M (1997) Dehydroergosterol structural organization in aqueous medium and in a model system of membranes. Biophys J 72:2226–2236

    Article  CAS  Google Scholar 

  56. Ghosh PB, Whitehouse MW (1968) 7-Chloro-4-nitrobenzo-2-oxa-1,3-diazole: a new fluorigenic reagent for amino acids and other amines. Biochem J 108:155–156

    CAS  Google Scholar 

  57. Monti JA, Christian ST, Shaw WA (1978) Synthesis and properties of a highly fluorescent derivative of phosphatidylethanolamine. J Lipid Res 19:222–228

    CAS  Google Scholar 

  58. Monti JA, Christian ST, Shaw WA, Finley WH (1977) Synthesis and properties of a fluorescent derivative of phosphatidylcholine. Life Sci 21:345–355

    Article  CAS  Google Scholar 

  59. Cairo CW, Key JA, Sadek CM (2010) Fluorescent small-molecule probes of biochemistry at the plasma membrane. Curr Opin Chem Biol 14:57–63

    Article  CAS  Google Scholar 

  60. Polyakova SM, Belov VN, Yan SF, Eggeling C, Ringemann C, Schwarzmann G, de Meijere A, Hell SW (2009) New GM1 ganglioside derivatives for selective single and double labelling of the natural glycosphingolipid skeleton. Eur J Org Chem 2009:5162–5177

    Article  Google Scholar 

  61. Uster PS, Pagano RE (1986) Resonance energy transfer microscopy: observations of membrane-bound fluorescent probes in model membranes and in living cells. J Cell Biol 103:1221–1234

    Article  CAS  Google Scholar 

  62. Pajk S, Garvas M, Štrancar J, Pečar S (2011) Nitroxide-fluorophore double probes: a potential tool for studying membrane heterogeneity by ESR and fluorescence. Org Biomol Chem 9:4150–4159

    Article  CAS  Google Scholar 

  63. Ryan TM, Griffin MDW, Bailey MF, Schuck P, Howlett GJ (2011) NBD-labeled phospholipid accelerates apolipoprotein C-II amyloid fibril formation but is not incorporated into mature fibrils. Biochemistry 50:9579–9586

    Article  CAS  Google Scholar 

Download references

Acknowledgments

Work in A.C.’s laboratory was supported by the Council of Scientific and Industrial Research and Department of Science and Technology, Government of India. S.H. thanks the Council of Scientific and Industrial Research for the award of a Senior Research Fellowship. A.C. is an Adjunct Professor at the Special Centre for Molecular Medicine of Jawaharlal Nehru University (New Delhi, India) and Indian Institute of Science Education and Research (Mohali, India) and Honorary Professor of the Jawaharlal Nehru Centre for Advanced Scientific Research (Bangalore, India). A.C. gratefully acknowledges J.C. Bose Fellowship (Dept. Science and Technology, Govt. of India). Some of the work described in this chapter was carried out by former members of A.C.’s research group whose contributions are gratefully acknowledged. We thank members of our laboratory for critically reading the manuscript. We dedicate this chapter to the memory of Prof. Richard E. Pagano for his seminal contribution in the development and application of NBD-labeled lipids in cell biology.

Author information

Authors and Affiliations

  1. Centre for Cellular and Molecular Biology, Council of Scientific and Industrial Research, Uppal Road, Hyderabad, 500 007, India

    Sourav Haldar & Amitabha Chattopadhyay

Authors
  1. Sourav Haldar
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Amitabha Chattopadhyay
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Amitabha Chattopadhyay .

Editor information

Editors and Affiliations

  1. Faculté de Pharmacie, Laboratoire de Biophotonique, Université de Strasbourg, route du Rhin 74, Illkirch, 67401, France

    Yves Mély

  2. Faculté de Pharmacie, Laboratoire de Biophotonique et, Université de Strasbourg, route du Rhin 74, Illkirch, 67401, France

    Guy Duportail

Rights and permissions

Reprints and permissions

Copyright information

© 2012 Springer-Verlag Berlin Heidelberg

About this chapter

Cite this chapter

Haldar, S., Chattopadhyay, A. (2012). Application of NBD-Labeled Lipids in Membrane and Cell Biology. In: Mély, Y., Duportail, G. (eds) Fluorescent Methods to Study Biological Membranes. Springer Series on Fluorescence, vol 13. Springer, Berlin, Heidelberg. https://doi.org/10.1007/4243_2012_43

Download citation

Publish with us

Policies and ethics

Search

Navigation