Bioassay Techniques for Drug Development

by Rahman; Atta-ur

• ISBN13: 9789058230515
• ISBN10: 9058230511
• Edition: 1st
• Format: Hardcover
• Copyright: 2001-10-04
• Publisher: CRC Press

Summary

The goal of an activity-directed isolation process is to isolate bioactive compounds which may provide structural leads of therapeutic importance. Whereas the traditional process of drug development is long and expensive, simple and rapid bioassays can serve as the starting point for drug discovery. This book presents a range of "bench top" bioassay techniques useful for natural product and pharmaceutical chemists involved in drug discovery and pharmacognosy. The contents detail a number of enzyme-based assays, cell-based functional bioassays and receptor radioligand binding assays along with detailed descriptions of each type. The majority of these bioassays are presented in a step-by-step format, so they could even be implemented by technical personnel with little background in microbiology, biochemistry or pharmacology.

Table of Contents

Preface	xi
General Introduction	1	(6)
Bench-Top and Primary Bioassay Screening	7	(100)
Toxicity Assays	9	(5)
Brine-shrimp lethality assay	9	(1)
Brine-shrimp microwell cytotoxicity assay	10	(2)
Crown gall tumor inhibition assay (potato disc antitumor assay)	12	(2)
Animal toxicity assay	14	(1)
Antimicrobial Assays	14	(11)
Antibacterial assays	16	(1)
Agar diffusion assay	16	(1)
Agar dilution assay	17	(1)
Microtitre plate method	18	(1)
Direct bioautobiography method	19	(3)
Antifungal assays	22	(1)
Agar tube dilution assay	22	(2)
Direct bioautobiography method	24	(1)
Antiviral and Anticancer Assays	25	(14)
Anti-HIV assay	25	(2)
Anticancer assays	27	(1)
In vitro anticancer screening	28	(9)
HCT cytotoxicity assay	37	(2)
Antimitotic Assay	39	(2)
Antimitotic assay using sea urchin eggs	39	(2)
Genotoxicity Assays	41	(7)
SOS chromtest for genotoxicity	41	(3)
Ames test for detecting carcinogens and mutagens	44	(3)
Antiphage activity assay	47	(1)
Assays for Control of Tropical Diseases	48	(17)
Antimalarial assays	48	(1)
96 well plate in vitro schizonticidal assay	49	(1)
Suppressive test of blood schizonticidal action (Peter's test)	50	(1)
Rane test of blood schizonticidal activity	51	(1)
Larvicidal assay	51	(1)
Molluscicidal assay	52	(1)
Piscicidal assay	53	(1)
Amoebicidal assay	54	(5)
Cercaricidal assay	59	(1)
Leishmanicidic activity	60	(1)
In vitro Leishmanicidal assay (promastigotes)	60	(2)
In vitro Leishmanicidal assay (amastigotes)	62	(2)
Leishmanicidal activity in vivo	64	(1)
Assays for Agrochemicals	65	(9)
Lemna minor for phytotoxicity and growth-stimulating assay	65	(2)
Contact toxicity insecticidal assay	67	(1)
Insect antifeedant assays	68	(1)
Leaf-choice assay	68	(1)
Artificial diet feeding bioassay	69	(3)
Nematicidal assay	72	(2)
Hepatotoxicity Assays	74	(8)
Antihepatotoxic activity using carbon tetrachloride-induced cytotoxicity	74	(2)
In vivo hepatoprotective assay	76	(2)
Antihepatotoxic activity using galactosamine induced cytotoxicity	78	(4)
Hypoglycemic/Antidiabetic Activity Assays	82	(5)
Antidiabetic activity assay on normal and alloxan-diabetic rabbits	82	(2)
Antidiabetic activity assay (GOD-PAP method)	84	(1)
Hypoglycemic activity assay	85	(2)
Diuretic Activity Assay	87	(1)
Anthelmintic Activity Assays	88	(2)
In vitro anthelmintic assay	88	(2)
In vivo anthelmintic assay	90	(1)
Antifertility/Anti-implantation Assays	90	(2)
Antifertility activity	90	(1)
Anti-implantation activity	91	(1)
In Vitro Assay for Platelet Aggregation	92	(2)
Anti-inflammatory Assay	94	(1)
Rat paw edema assay	94	(1)
Immunomodulating Assay	95	(1)
Hemolytic plaque assay	95	(1)
Antiepileptic (Anticonvulsant) Assay	96	(4)
Pilocarpine model	97	(3)
Analgesic Assays	100	(1)
Writhing syndrome method	100	(1)
Hot plate method	100	(1)
Gastroprotective/Antiulcer Assays	101	(2)
Gastroprotective assay	101	(1)
Anti-ulcerogenic assay-1	102	(1)
Anti-ulcerogenic assay-2	103	(1)
Radiolabelling Bioassays	103	(3)
Mitogenic activity assay	103	(3)
Anti-emetic Assay	106	(1)
High-Throughput Screening	107	(112)
Introduction	109	(5)
Characteristics of an ideal screening assay for screening natural product extracts	109	(1)
Biochemical or cell based assays	110	(1)
Assay format	110	(1)
Reagent source	111	(1)
Assay signal-to-noise ratio	112	(1)
Assay incubation	112	(1)
Assay manipulations	113	(1)
DMSO compatibility	113	(1)
Screening source material compatibility	113	(1)
Enzyme Assays	114	(38)
Introduction	114	(1)
Considerations for development of enzyme assays	114	(1)
Selection of enzyme source	115	(1)
Selection of enzyme concentration	116	(1)
Selection of substrate and substrate concentration	116	(5)
Selection of cofactor concentrations	121	(1)
Selection of assay endpoint	122	(1)
Selection of assay pH	122	(1)
Selection of assay temperature	123	(1)
DMSO compatibility	123	(1)
Extract compatibility	123	(1)
Inhibitor profiles	123	(1)
Competitive inhibition	124	(1)
Uncompetitive inhibition	125	(1)
Noncompetitive inhibition	125	(1)
Development of a high throughput enzyme assay for natural product screening: an example	126	(7)
Some common in vitro enzyme-based bioassays	133	(1)
Protease inhibition assays	133	(4)
Tyrosinase inhibition assay	137	(2)
Hyaluronidase inhibition assay	139	(3)
Acetylcholinesterase inhibition assay	142	(1)
Spectrophotometric assay	143	(2)
β-Lactamase inhibition assay	145	(1)
Spectrophotometric assay	146	(2)
5-Lipoxygenase inhibition assay	148	(1)
Spectrophotometric assay	149	(3)
Cell-based Receptor Functional Assays	152	(27)
Introduction	152	(1)
Utility of receptor functional assays	152	(1)
Functional assays for G-protein coupled receptors	152	(1)
Gs and Gi coupled receptor signal transduction pathways	152	(3)
High throughput functional assays for Gs and Gi coupled receptors	155	(1)
Common cell-based receptor functional assays	155	(1)
SPA cAMP assay	155	(1)
FlashPlate cAMP assay	156	(1)
ELISA-based cAMP assays	157	(3)
cAMP response element reporter gene assay	160	(1)
Gq-coupled receptor signal transduction pathways	161	(1)
Introduction	161	(3)
High throughput assays for Gq-coupled receptors	164	(1)
High throughput functional assays for ligand-gated ion channels	165	(2)
Receptor protein tyrosine kinase functional assays	167	(1)
Introduction	167	(1)
RPTK-associated signal transduction pathways	167	(2)
High throughput functional assays for RPTK'S	169	(1)
Intracellular calcium assay	169	(1)
Proliferation assays	170	(3)
Cytokine receptor functional assays	173	(1)
Cytokine receptor signal transduction pathways	173	(1)
Functional assays for cytokine receptors	173	(1)
Mixed receptor functional assay	174	(1)
Melanophore functional receptor assay technology	174	(4)
Receptor selection and amplification functional receptor assay technology	178	(1)
Radioligand Binding Assays	179	(26)
Introduction	179	(1)
Important considerations for developing radioligand binding assays	180	(1)
Receptor source and assay concentration	180	(1)
Nonrecombinant human cell lines	180	(1)
Membrane preparations from nonrecombinant cells and tissues	181	(1)
Recombinant receptor containing cells or membranes	181	(1)
Selection of radioligand and assay concentration	182	(2)
Selection of assay incubation conditions	184	(1)
Selection of assay buffer	184	(1)
Selection of assay volume	184	(1)
Selection of a methodology for separation of free and bound radioligand	185	(1)
Steps in developing and validating radioligand binding	185	(1)
Steps in developing and validating radioligand binding assays	185	(1)
Cell membrane linearity evaluation	186	(1)
Association and dissociation time courses	187	(5)
Saturation isotherm evaluation	192	(2)
Competition studies	194	(4)
Alternative radioligand binding assays	198	(1)
FlashPlate assays	198	(2)
Scintillation proximity assays	200	(1)
ScintiStrip plate assays	201	(2)
Fluorescence based receptor binding	203	(1)
Time resolved fluorescence	203	(1)
Fluorescence polarization	203	(2)
References	205	(14)
Index	219