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HUMAN GENETICS

A LABORATORY MANUAL FOR HUMAN BLOOD ANALYSIS

M.K. BHASIN (University of Delhi, Delhi, India) AND

S.M.S. CHAHAL (Punjabi University Patiala, Punjab, India)

With a Foreword by: S. C. TIWARI

First Printed: 1996 • Reprinted: 2013 • Pages: 351 • Size: 180 x 240 mm •

ISBN 81-85264-12-0 •Binding : Hard •

Price: US $ 45/- Rs. 1450/-

ANTHROPOMETRY

The “Laboratory Manual for Human Blood Analysis” is a unique textbook. It is “must be” for all laboratories, in which the genetic blood group systems of man are analysed. This “Laboratory Manual” does not offer only excellent possibilities to the beginner to become familiar with the multifaced experimental and statistical methods employed in serological population genetics, but is also an excellent laboratory reference book for the advanced scholar. That is why one wish a broad distribution of this book. There is no doubt that in future the “Laboratory Manual” will become the Standard Text and Reference Book to all those who in research or in practice wish to work or are already working in the field of the genetic markers of the human blood. —ANTHROPOLOGISCHER ANZEIGER (Germany)

Human blood studies, both theoretical and practical, are an integral part of curricula of varied disciplines - Medical Sciences, Biological Anthropology, Zoology, Forensic Science, Genetics, Life Sciences, Human Biology, but there seems to be great paucity of books covering laboratory techniques. The present book covers the latest techniques, giving their description and methods of execution in lucid terms. The book has thirteen chapters:

CONTENTS

Foreword

Preface

List of Figures

List of Tables

  1. BLOOD

    1. Characteristics of Blood

    2. Collection of Blood

      1. Equipment,

      2. Collection of Blood from Finger-Tip,

      3. Capillary Tube Method,

      4. Venipuncture Method

    3. Preparation of Blood

    4. Anticoagulants

    5. Isotonic Solution

      6. Appendix

    6. Basic Equipments and Apparatus

    7. Abbreviations

    8. Equivalents of Measures and Weights

  2. TECHNIQUE FOR BLOOD CELL EXAMINATIONS

    1. Complete Blood Count

      1. White Cell Count,

      2. Red Cell Count,

      3. Haemoglobin Determination,

      4. Differential White Cell Count and Stained Red Cell Examination,

        1. Differential White Cell Count,

        2. Stained Red Cell Examination)

    2. Sedimentation Rate

    3. Tests to Study Types of Anaemia

      1. Haematocrit (Hct) Reading or Packed Cell Volume (PCV),

      2. Reticulocyte Count,

      3. Osmotic Fragility Test,

      4. Mean Corpuscular Values,

      5. Red Cell Indices)

      4. Appendix

    4. Haemoglobin Nomogram

  3. Haemoglobins

    1. Normal Haemoglobins

    2. Abnormal haemoglobins

    3. Techniques

      1. Haematological Studies,

      2. Preparation of Red Blood Cell Haemolysate,

      3. Electrophoretic Techniques,

      4. Antenatal Diagnosis of Haemoglobinopathies,

      5. Sickle Cell Examination,

      6. Determination of Haemoglobin A2,

      7. Determination of Foetal Haemoglobin (Hb F)

      4. Appendix

    4. Haemoglobins

      1. Haemoglobin S,

      2. Haemoglobin C,

      3. Haemoglobin D,

      4. Haemoglobin E,

      5. Hereditary Persistence of Foetal haemoglobin,

      6. Thalassaemia,

      7. Other Haemoglobin Variants

    5. List of Haemoglobin Variants

    6. Guidelines for Human Gene Nomenclature (ISGN, 1987)

      1. Gene and Allele Terminology,

      2. Genotype Terminology,

      3. Phenotype Terminology,

      4. Haemoglobin Nomenclature)

  4. Glucose-6-Phosphate Dehydrogenase

    1. Introduction

    2. Quantitative Determination

    3. Molecular Forms

    4. Techniques

      1. Screening Procedures,

      2. Quantitative Techniques Reflecting G6PD Activity,

      3. Electrophoretic Characterisation of G6PD Variants,

      4. Partial Purification of Human Red Cell (G6PD),

      5. Determination of Substrate Affinity or Michaelis Constant (Km),

      6. Utilization of Substrate Analogues,

      7. Thermostability,

      8. Other Kinetic Measurements,

      9. Other Means of Characterising G6PD

      5. Appendix

    5. List of G6PD Variants

  5. Techniques in Blood Grouping

    1. Apparatus and Chemicals

    2. Precautions

      1. Cleaning of Dirty Glassware

    3. Designation of Agglutination Reaction

    4. Blood Group Terminology

    5. The ABO Blood Group System

      1. The ABO Blood Groups,

      2. Method for Detecting Subgroups of A and AB,

      3. The Bombay (Oh) Phenotype

    6. The MNSs Blood Group System

      1. The MN Blood Groups,

      2. The MNS Blood Groups)

    7. The P Blood Group System

    8. The Rhesus Blood Group System (

      1. Anti-D (Anti-Rh 0) Test Method),

      2. Du Test Method,

      3. Rh Complete Sera (IgM) (Saline Agglutinating),

      4. Rh Incomplete Sera (IgG) (Albumin Agglutinating)

    9. Secretors and Non-Secretors (The ABH Secretion System)

    10. The Lewis System

    11. Blood Grouping By Indirect Antiglobulin Test (Coombs Test)

      1. The Lutheran Blood Group System,

      2. The Kell Blood Group System,

        1. The Sutter Groups,

      3. The Duffy Blood Group System,

      4. The Kidd Blood Group System,

      5. The Diego Blood Group System,

      6. The Colton Blood Group System,

      7. The Xg Blood Group System)

    12. Anti-Human Globulin Serum (For Direct or Indirect Antiglobulin/Coombs Tests)

    13. Blood Grouping from Bloodstains, Fragments of Muscle, Skin etc. and Lysed Blood Samples

      1. The ABO Blood Groups,

      2. The MN Blood Groups,

      3. The Rh Blood Groups,

      4. Grouping Tests on Saliva

      14. Appendix

    14. Blood as a Forensic Indicator

  6. Techniques in Human Leukocyte Grouping

    1. The HLA System

    2. Techniques

      1. Serological Methods,

      2. Data Analysis,

      3. Reasons for False Reactions,

      4. Quality Control

    7. Appendix

  7. Techniques in Immunoglobulin Typing

    1. Immunoglobulin Plasma Protein Allotypes: The Gm, Km (Inv) and Am Systems

    2. Techniques in Immunoglobulin Typing: Gamma Globulin Factors (Gm) and Kappa Markers (Km)

      1. Agglutination-Inhibition Tests

  8. TECHNIQUES IN ELECTROPHORESIS

    1. Electrophoresis

      1. Apparatus for Electrophoresis,

      2. Applications of Electrophoresis,

      3. Starch Gel Electrophoresis,

      4. Agarose Gel Electrophoresis,

      5. Cellulose Acetate Electrophoresis,

      6. Polyacrylamide Gel Electrophoresis,

      7. Block Electrophoresis,

      8. Isolectric Focusing,

      9. Disc Electrophoresis,

      10. Protein Blotting,

      11. Pulsed Field Gel Electrophoresis

    2. Immunological Techniques

      1. Immunoelectrophoresis,

      2. Immunodiffusion

      3. Appendix

    3. Guidelines for Human Gene Nomenclature (ISGN, 1987)

      1. Enzyme and Protein Nomenclature

  9. SERUM PROTEIN POLYMORPHISMS

    1. Introduction

    2. Protein Polymorphisms

      1. Haptoglobin (Hp) System,

      2. Serum Transferrin (Tf) System,

      3. Group Specific Component (Gc) System,

      4. Complement Component 3 (C3) System,

      5. Properdin Factor B (Bf) System

  10. RED CELL ENZYME POLYMORPHISMS

    1. Introduction

    2. Enzyme Polymorphisms

      1. 6-Phosphogluconate Dehydrogenase (6-PGD) System (E.C. 1.1.1.44),

      2. Adenylate Kinase (AK) system (E.C. 2.7.4.3),

      3. Phosphoglucomutase Locus 1 (PGM1) System (E.C. 5.4.2.8 earlier E.C. 2.7.5.1),

      4. Acid Phosphatase (ACP) System (E.C. 3.1.3.2),

      5. Adenosine Deaminase (ADA) System (E.C. 3.5.4.4),

      6. Esterase D (EsD) System (E.C. 3.1.1.1.),

      7. Glyoxalase I (GLO I) System (E.C. 4.4.1.5),

      8. Phosphohexose Isomerase (PHI) System (E.C. 5.3.1.9),

      9. Glutamate Pyruvate Transaminase (GPT) System (E.C. 2.6.1.2.)

      3. Appendix

    3. Scheme of Classification and Numbering of Eyzymes

  11. DNA POLYMORPHISMS

    1. Introduction

    2. Reagents, Apparatus and Equipments

    3. Techniques

      1. Isolating Pure DNA from Human Cell (Specimen Collection and Preparation for Analysis),

        1. Cell Preparation,

        2. Extraction of DNA

        3. Determination of DNA Concentration by Spectrophotometry,

      2. Restriction Enzyme Digestion,

      3. Agarose Gel Electrophoresis,

      4. Southern Transfer,

      5. DNA Hybridization,

      6. Colour Development and Autoradiography,

        1. Colour Development,

        2. Autoradiography,

        3. Interpretation of Photograph or Autorads,

      7. Cell Preparation and Extraction of DNA From Forensic Exhibits

    12. Appendix

  12. OTHER GENETIC TRAITS

    1. Colour Blindness

      1. Technique

    2. Taste Testing with Phenylthiocarbamide (PTC)

      1. Technique,

      2. Allele Frequency Estimations

    3. Ability to Smell Hydrogen Cyanide

    4. Human Cerumen Type

      1. Technique,

      2. Allele Frequency Estimations

  13. STATISTICAL ANALYSIS

    1. Allele Frequency Calculations

      1. The ABO System,

        1. Allele Frequencies when Anti-A and Anti-B have been used,

        2. Allele Frequencies when Anti-A, Anti-B and Anti-A1 have been used,

      2. The MNSs System,

        1. Allele Frequencies when Anti-M and Anti-N have been used,

        2. Haplotype Frequencies when Anti-M, Anti-N, and Anti-S have been used,

        3. Haplotype Frequencies when Anti-M, Anti-N, Anti-S and Anti-s have been used,

      3. The Rh System,

        1. Allele Frequencies when Anti-d has been used,

        2. Haplotype Frequencies when Anti-C (rh’), Anti-D (Rh0), anti-E (rh”), Anti-c (hr’) and Anti-E (hr”) have been used,

      4. The Kidd (Jk) System,

        1. Allele Frequencies when only Anti-Jka has been used,

        2. Allele Frequencies when both Anti-Jka and Anti-Jkb have been used,

      5. Acid Phosphatase (ACP) System,

      6. The Gm (1,2,5) System,

      7. X-Linked Systems,

        1. The Xg System,

        2. Glucose-6 Phosphate Dehydrogenase (G6PD) System

    2. Estimation of Measures of Genetic Variation and Genetic Distance

      1. Heterozygosity,

      2. Genetic Differentiation,

        1. Wahlund’s Variance,

        2. Nei’s Gene Diversity Analysis,

      3. Genetic Distance,

        1. Steps of Computation of Nei’s Standard Genetic Distance

    3. Calculations Pertaining to Forensic Applications

      1. Measures of Individualization and Discrimination Potential,

      2. Measures of Paternity Exclusion,

        1. Measure of Paternity Exclusion Based on Single System,

        2. Measure of Paternity Exclusion Based on More than One System)

      4. GLOSSARY

      APPENDIX

      1. List of Chemicals, Their Abbreviations, Molecular Weights and Specifications

      2. Sources of Reagents and Equipments

      BIBLIOGRAPHY

      INDEX

Almost all recent researches in serology and haemotology have been consulted for preparing this manual.

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