黑料网

Uncharted Territory: The Specific Detection and Quantification of Lipids

Rupak Doshi^*

Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego , 9500 Gilman Drive, La Jolla 92093, USA

*Corresponding Author:

Rupak Doshi
Skaggs School of Pharmacy and Pharmaceutical Sciences
University of California, San Diego
9500 Gilman Drive, La Jolla 92093, USA
Tel: 44-1223-334-073
E-mail: rudoshi@ucsd.edu

Received date: April 29, 2013; Accepted date: April 30, 2013; Published date: May 02, 2013

Citation: Doshi R (2013) Uncharted Territory: The Specific Detection and Quantification of Lipids. J Anal Bioanal Tech 4:e114. doi: 10.4172/2155-9872.1000e114

Copyright: © 2013 Doshi R. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Visit for more related articles at Journal of Analytical & Bioanalytical Techniques

Few biological reagents have been more useful in unravelling the mysteries of biology than antibodies. Highly specific antibodies generated against particular antigens of interest have led to the detection and quantification of numerous targets. Today, antibodies are routinely used as sensors/probes in biochemistry, cell imaging, structural biology etc. to achieve various aims [1].

The process of generating antibodies against targets of interest has been highly successful against proteinacious antigens [2]. While this approach has been incredibly useful in studying protein expression, trafficking, signal transduction pathways etc., it neglects the rest of cellular components from analysis. Recent years have seen some success in the discovery of antibodies against non-proteinacious targets such as glycans, sugars etc., [3]. In comparison, success has been almost impossible to come by in the case of antigens that are “greasy” or hydrophobic in nature, for e.g. lipids and oils.

Hydrophobic entities play a critical role in the cell [4]. Lipids serve as membranes/borders around the cell, >70% of which is water. Lipids also border the intracellular organelles, such as mitochondria, endoplasmic reticulum and the nucleus [4]. In recent years, another important role has emerged for lipids in signal transduction [4]. Many plants and animals use cell-synthesized oils for various functions, most notably being intermediates of important metabolic pathways, and for defence. Plants also produce waxes and related lipids that form the cuticle around the cell [5].

With the above critical roles of hydrophobic compounds in biology, the absence of specific antibodies for detection and quantification is striking. There are two major methods to generate antibodies against a target antigen; (1) In vivo, using animals [6], (2) In vitro, using display technologies [7]. Lipids and related hydrophobic compounds are found in all living organisms and so the in vivo approach may not be ideal, because for it to work, the substance needs to elicit a strong immune response to a “foreign” antigen. Over the last few years, several in vitro display technologies have emerged to select and evolve highaffinity antibodies against antigens of interest. The process involves 3 major steps [7]:

a. generating a large antibody library containing several random sequences.

b. exposing the library to the antigen of interest, while maintaining a link between an antibody with its coding sequence; such a genotypephenotype linkage is achieved through various “display” methods such as on E. coli or yeast whole cells, bacteriophages, ribosomes, mRNA/ cDNA etc.

c. affinity maturation of the selected antibodies to increase the binding affinity to the antigen through rational and/or randomized mutagenesis.

There are several problems associated with lipidic antigens. These include but are not limited to the availability of large amounts of purified sample, in vitro immobilization on a solid surface, presence of multiple epitopes on the molecular surface, repeated units/polymerisation, the hydrophobic chemical nature which favours non-specific hydrophobic interactions/associations as opposed to specific ionic interactions etc. A concerted effort through the help and collaboration of chemists, physicists, engineers and of course, biologists will be required in the coming years, in order to navigate through these problems.

Furthermore, the clinical drug development pharmaceutical companies must also show interest in this area [8]. Antibodies against extremely hydrophobic drugs will allow specific transport assays to be conducted; these data are critical towards assessing the bioavailability, tissue distribution and body clearance of drugs in clinical trials. Regulatory bodies such as the USFDA require direct answers to whether drug candidates are transport substrates of multidrug efflux transporters like ABCB1, BCRP etc., [9]. As opposed to the current indirect practices of using fluorescent compounds to measure transport of a drug candidate, one should be able to measure the trans-membrane movement of the drug candidate itself directly, using antibodies as drug-sensors.

Another emerging area where anti-lipid antibodies will make a huge impact is biofuels [10]. Several algae, yeast, cyanobacteria, diatoms and bacteria are being currently engineered to over-produce biofuel molecules. However, the measurement of these compounds remains a big challenge [10]. At present, costly mass spectrometry methods are required to quantify the production in massive screens that involve many tens or hundreds of samples. Rapid antibody based quantification methods could really aid biofuels industry [11] which is poised to constitute ~5% of the global fuel needs.

The availability of highly specific antibodies against lipids, drugs and oils will not only be a tremendous boost to cell biology research in all 5 kingdoms of life, but will also aid the pharmaceutical and environmental/energy industries in their efforts. Thus, there is little doubt that we will see a large amount of funding and effort put into this area in the coming few years.

References

1. Maynard J, Georgiou G (2000)
2. Konthur Z, Hust M, Dübel S (2005)
3. Dotan N, Altstock RT, Schwarz M, Dukler A (2006)
4. Cullis PR, de Kruijff B (1979)
5. Samuels L, Kunst L, Jetter R (2008)
6. Kellermann SA, Green LL (2002)
7. Kretzschmar T, von Rüden T (2002)
8. Zhang L, Strong JM, Qiu W, Lesko LJ, Huang SM (2006)
9. International Transporter Consortium, Giacomini KM, Huang SM, Tweedie DJ, Benet LZ, et al. (2010)
10. Pienkos PT, Darzins AL (2009)
11. DeMartini JD, Pattathil S, Utku Avci U, Szekalski K, Mazumder K, et al. (2011)