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Section 3 : Immunologic Methods and Testing

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Abstract:

This section talks about immunologic methods, testing and discusses about sample collection, processing, and handling. The section also discusses the laboratory procedures involved in clinical immunology. Sufficient detail is provided to enable the reader to understand the basic theory behind each procedure, the types of specimens required, and the possible problems and pitfalls. The processing and handling of samples are discussed to emphasize that correct handling of samples is as important as having the correct sample. The methods used in the clinical immunology laboratory have evolved over many decades. Although the initial serologic techniques were developed in the late 19th century, the major growth of this discipline has occurred in the last 30 or so years, following the development of RID, IIF, RIA, and EIA. Examples of potentially infectious material include blood and blood products, semen, vaginal fluids, CSF, pleural fluid, pericardial fluid, peritoneal fluid, amniotic fluid, and any unfixed human tissues or organs. Specimens should be collected in the correct containers; in most cases, this is dictated by the type of anticoagulant needed. Any special handling or storage requirements for specimens, such as stat transportation to the laboratory for total hemolytic complement (CH) determinations, should be noted on the order sheet. Although most specimens are safe at room temperature for a short time, perhaps 30 to 60 min, some specimens must receive special handling. These include peripheral blood and fluids for leukocyte phenotyping and function and blood drawn for complement and/or cryoglobulin assays.

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3

Key Concept Ranking

Complement System
0.6152139
Cytotoxic T Cell
0.46756482
Clinical Immunology
0.45893368
White Blood Cells
0.4367687
Agarose Gel Electrophoresis
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0.6152139
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Figures

Image of Figure III.1
Figure III.1

SPE and UPE. (a) Normal SPE; (b) polyclonal gammopathy, serum; (c) monoclonal gammopathy, serum; (d) hypogammaglobulinemia, serum; (e) monoclonal gammopathy, urine; (f) proteinuria, glomerular; (g) proteinuria, tubular; (h) proteinuria, mixed glomerular and tubular.

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3
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Image of Figure III.2
Figure III.2

Hemoglobin electrophoresis. ( a) Alkaline gel: patient 1 (42 years old; male) shows Hb A and Hb F; patients 2 (31 years old, female) and 3 (18 years old, female) show Hb A and Hb S. (b) Acid gel: further resolution of samples 1, 2, and 3 at acid pH shows that patient 1 has 77% A and 21% F; both patients 2 and 3 have 60% A and 38% S. C, Hb C; S, Hb S; F, Hb F; A, Hb A.

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3
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Image of Figure III.3
Figure III.3

Immunofixation. (a) SPE of two sera that contain monoclonal immunoglobulins: patient 4 serum has a single monoclonal component in the β region; patient 5 serum has two bands in the γ region and possibly a third band in the β region. (b) Immunofixation of the sera: patient 4 serum contains an IgA λ monoclonal immunoglobulin; patient 5 serum contains two IgG κ monoclonal proteins and an IgA λ monoclonal protein.

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3
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Image of Figure III.4
Figure III.4

Examples of oligoclonal bands in CSF. Matched serum samples were diluted to the same IgG concentration as the CSF sample. (1) Positive control (commercial); (2) CSF, patient 6 (positive for oligoclonal bands); (3) serum, patient 6; (4) pI markers; (5) CSF, patient 7 (positive for oligoclonal bands); (6) serum, patient 7; (7) negative control. Note the cathodic IgG bands seen in the CSF of patients 6 and 7, but not in the matched serum samples.

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3
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Image of Figure III.5
Figure III.5

Algorithm for analysis of proteins in serum and urine. *, computerized database of previous University of Virginia SPE and UPE results; QIgs, κ, λ, quantitation of IgG, IgA, IgM, and κ and λ light chains; IFE, immunofixation; TP, total protein; α, β, γ, alpha-2, beta, and gamma fractions of serum by SPE. Modified from Normansell (1994) with permission.

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3
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Image of Figure III.6
Figure III.6

Detection of antibodies to HIV-1 by IB. Strips: (1) strong positive control; (2) weak positive control; (3) negative control; (4) patient 8, positive (gp160, gp120, p65, p55, p51, gp41–43, p32, p24, p18, p15); (5) patient 9, positive (gp160, gp120, p65, p55, p51, gp41–43, p32, p24, p18, p15); (6) patient 10, negative; (7) patient 11, positive (gp160, gp120, p65, p55, p51, gp41–43, p32, p24, p18).

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3
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Image of Figure III.7
Figure III.7

HLA allele distribution at the HLA locus onchromosome 6. From Normansell (1994) with permission.

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3
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Image of Figure III.8
Figure III.8

Amplification procedures in current use. (a) PCR. The first four cycles are shown; the dominant product is the sequence between the two primers (these are indicated by the small vertical bars on the initial DNA molecule). (b) LCR. The first cycle is shown. (c) bDNA procedure. Capture probes anchor target-specific probes to the solid matrix. Target-specific probes also bind the amplifier molecules, which are branched DNA molecules. A large number of small enzyme-labelled probes are bound to the DNA molecules for reaction with the chemiluminescent substrate.

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3
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References

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Tables

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Table III.1

Components of SPE fractions

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3
Generic image for table
Table III.2

HLA alleles

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3
Generic image for table
Table III.3

Types of Vacutainer tubesa and order of draw

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3
Generic image for table
Table III.4

Specimens required for commonly requested immunology tests

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3
Generic image for table
Table III.5

Serum protein reference intervals for immunoglobulins

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3
Generic image for table
Table III.6

Serum protein reference intervals for proteins other than IgG, IgA, and IgM

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3
Generic image for table
Table III.7

Reference intervals for lymphocyte subsets

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3
Generic image for table
Table III.8

Reference intervals for CBC and white blood cell differential used at the University of Virginia

Citation: Folds J, Normansell D. 1999. Immunologic Methods and Testing, p 187-246. In Pocket Guide to Clinical Immunology. ASM Press, Washington, DC. doi: 10.1128/9781555818197.ch3

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