About the Book

• Meant for MBBS students.
• Based on the Kathmandu University School of Medical Sciences curriculum for MBBS.
• The content of the manual compliments with what is being taught in the theory classes.
• Clinically relevant questions about experiments are included at the end of the chapter.
• Includes exercises on protein and DNA electrophoresis.

About the Author

Arti S Pandey PhD Arun Pandey MSc Naveen K Shreevastava MSc Durga P Neupane MSc

Table of Contents:
MBBS I Experiment 1: Use of Micropipettes 3 • Introduction 3 • Use 3 • Procedure for micropipetting 4 • Checking your pipetting skill 5 Experiment 2: Buffers and pH 7 • Introduction 7 • pH meter 8 Experiment 3: Qualitative tests for Carbohydrates 11 • Introduction 11 • Reactions of monosaccharides, disaccharides and polysaccharides 11 Experiment 4: Qualitative Tests for Proteins 20 • Introduction 20 • Biuret reaction (general test for protein) 20 • Heat coagulation test 21 • Precipitation by salt solution (ammonium sulfate) 21 Experiment 5: Separation of Serum Proteins by Gel Electrophoresis 25 • Introduction 25 • Principle 26 • Quantitation 27 • Hyperproteinemias 28 • Hypoproteinemias 28 Experiment 6: Principles of Colorimetry 32 • Introduction 32 • The photoelectric colorimeter 33• Verification of Beer-Lambert's law using bromophenol blue 34 Experiment 7: Estimation of Serum Total Protein and Determination of Albumin to Globulin Ratio 38 • Introduction 38 • Principle 38 • Reagents 38 • Procedure 39 • Observation 39 • Calculation 39 • Normal range (adults) 40 Experiment 8: Genomic DNA Isolation, Amplification of the D1S80 Locus and DNA-Gel Electrophoresis 42 • Introduction 42 • Materials and methods 43 • The D1S80 locus 44 • Gel electrophoresis and DNA fragment analysis 46 Experiment 9: Estimation of Blood Glucose by GOD-POD Method 49 • Introduction 49 • Principle 49 • Specimen collection 49 • Reagents 50 • Sample collection 50 • Procedure 50 • Observation 50 • Normal range (serum/plasma glucose) 51 • Comment on the result 51 Experiment 10: Estimation of Serum Total Cholesterol by Modified Zak’s Method 53 • Introduction 53 • Principle 53 • Reagents 54 viii Biochemistry Laboratory Manual for MBBS (I and II) • Procedure 54 • Observation 55 • Calculation 55 • Normal range for serum cholesterol (recommended by NCEP) 55 • Comment on the result 55 Experiment 11: Estimation of Serum Triglycerides 57 • Introduction 57 • Principle 57 • Procedure 57 • Observations 58 • Calculation 58 • Normal values: (A/C to NCEP-ATP II) 58 • Comment on the result 59 Experiment 12: Estimation of Serum Uric Acid by Caraway’s Method 61 • Introduction 61 • Principle 61 • Reagents 62 • Procedure 62 • Observations 62 • Calculation 63 • Normal range 63 • Comment on the result 63 MBBS II Experiment 13: Determination of Serum Amylase Activity by Amyloclastic Method 67 • Introduction 67 • Principle 67 • Reagents 68 • Procedure 68 • Normal range 69 • Comment on the result 69 14: Estimation of Serum Aspartate Transaminase (AST) Activity 71 • Introduction 71 • Principle 71 • Reagents 72 • Procedure 72 • Observations 72 • Normal range 73 • Comment on the result 73 Experiment 15: Estimation of Serum Alanine Transaminase (ALT) Activity 75 • Introduction 75 • Principle 76 • Reagents 76 • Procedure 76 • Observations 77 • Calculation 77 • Normal range 77 • Comment on the result 78 Experiment 16: Estimation of Serum Bilirubin by Modified Jendrassik/Grof Method 80 • Introduction 80 • Principle 81 • Reagents 81 • Procedure 81 • Observations 82 • Calculation 82 • Normal range 83 • Comment on the result 83 Experiment 17: Estimation of Serum Urea by Diacetyl Monoxime (DAM) Method 87 • Introduction 87 • Principle 88 • Reagents 88 x Biochemistry Laboratory Manual for MBBS (I and II) • Procedure 88 • Observation 89 • Calculation 89 • Normal range 89 • Comment on the result 90 Experiment 18: Estimation of Serum Creatinine by Jaffe’s Alkaline Picrate Method 91 • Introduction 91 • Principle 91 • Reagents 92 • Procedure 92 • Observation 92 • Calculation 93 • Normal range 93 • Comment on the result 93 • True creatinine 93 Experiment 19: Urinary Creatinine (Jaffe’s Method) and Creatinine Clearance 95 • Introduction 95 • Principle 95 • Procedure 95 • Observation 96 • Calculation 96 • Normal range 97 • Comment on the result 97 • Normal range 98 • Comment on the result 98 Experiment 20: Urinalysis 101 • Introduction 101 • Physical examination 101 • Chemical examination 102 • Abnormal chemical constituents 103 Contents xi Experiment 21: Urinalysis (Abnormal Constituents) 108 • Physical examination of urine 108 • Chemical examination 108 • Result 109 • Interpretation 109 Experiment 22: Oral Glucose Tolerance Test (OGTT): Principle and Interpretation 112 • Introduction 112 • Preparatory phase 112 • Procedure 113 Experiment 23: Molecular Diagnosis: Example Sickle Cell Anemia using ASO and RFLP 116 • Introduction 116 • Sickle cell anemia 116 Reference Biochemical Values 121 Reagents 122