Chapter 11: Serology Techniques: Past, Current, and Future

Forensic serology

Forensic serology

is the component of forensic biology that deals with the examination and identification of biological evidence.

Short Tandem Repeats (STR)

The most commonly utilized forensic DNA analysis used for human identification.

Y-chromosomal STR analysis

is often utilized for the investigation of sexual assault crimes.

Mitochondrial DNA analysis

is used for the identification of human remains.

presumptive and confirmatory assays

The identification of bodily fluids can be carried out using ______ to identify the type of bodily fluid in question.

Positive assay

indicates the possibility of the presence of the bodily fluid in question.

Negative assay

indicates that the questioned bodily fluid is absent.

Confirmatory Assays

These assays are utilized to identify bodily fluids with higher certainty than presumptive assays.

Primary Binding Assays

It involves the initial binding between a single epitope of an antigen and a single binding site of an antibody. They are very sensitive.

Secondary Binding Assays

They are less sensitive and easier to perform than primary binding assays.

Precipitation-based assays

________ have been used for species identification.

Agglutination-based assays

______ are more sensitive. It detects antigens located on the surface of cells or carriers, are normally applied to blood group typing.

Enzyme-Linked Immunosorbent Assay (ELISA)

It is an immuno-enzyme assay that can be used to detect and measure the antibody or antigen in question.

Antibody-sandwich ELISA

The most common ELISA that is used in forensic serology.

Immunodiffusion

A passive method in which an antigen or an antibody or both are allowed to diffuse and therefore a gradient, from low to high concentration, is established for an antigen or an antibody or both.

Single Immunodiffusion

A concentration gradient is established for either an antigen or an antibody.

Double Immunodiffusion

A concentration gradient is established for both an antigen and an antibody.

Immunoelectrophoresis (IEP)

This technique uses electrophoresis to separate the antigen mixture prior to immunodiffusion.

Crossed Immunoelectrophoresis

Also known as two-dimensional IEP, is a modification of IEP.

Rocket Immunoelectrophoresis

An antibody-containing agarose gel is used. The antigen is loaded into the well.

Crossed-Over Immunoelectrophoresis

This technique is also known as counterimmunoelectrophoresis (CIE).

Agglutination reactions

can be used as forensic serological assays such as for blood group typing and menstrual blood identification.

Agglutination assays

are qualitative, indicating the absence or presence of antigens or antibodies.

Semi-quantitative assays

results can be obtained by titration.

Direct agglutination assays

involve reactions in which an antibody interacts with antigens originally located on cell surfaces.

hemagglutination reaction

In a ______, an antibody binds to the antigens located on erythrocytes.

Agglutination inhibition assays

The presence of an antigen in question is indirectly detected.

Passive agglutination assays

The antigen is coated on the surface of carrier cells such as tannic acid–treated sheep erythrocytes.

CpG dinucleotide sequences

In the eukaryotic genome, methylation occurs at the cytosine residues commonly in the _________ of both DNA strands.

phosphodiester

The “p” in CpG refers to the _______ bond between cytosine and guanine.

Cytosine methylation

is carried out in vivo by methyltransferase.

Genomic Loci

Also known as tissue-specific differentially methylated regions (TDMRs).

Methylation-sensitive restriction enzyme digestion polymerase chain reaction (MSRE-PCR)

can be used for the rapid detection of DNA methylation.

Bisulfite sequencing

The genomic DNA is treated with sodium bisulfite, which catalyzes the hydrolytic deamination of unmethylated cytosines.

Methyl-DNA immunoprecipitation (MeDIP)

is a useful method for isolating methylated DNA.

Messenger RNA-Based Assays

can potentially be used for wound age estimation and the age of biological stains.

Reference genes

Are constitutively expressed housekeeping genes are utilized as internal controls.

MicroRNA-Based Assays

The biological function of microRNAs (miRNAs) is to regulate gene expression.

Mass spectrometry (MS)

is a susceptible and rapid technique for protein identification from complex biological samples.

Mass spectrometer

An analytical technique to identify a molecule by measuring the ratio of the mass (m) to the charge (z) of a charged molecule.

Ion source

Converts analytes into gaseous phase ions through ionization.

Ionization

A process used by gaining a positive or a negative charge from a neutral species.

Mass Analyzer

A part of a mass spectrometer where ions are accelerated under electric fields and separated based on their m/z ratio.

Biomarker proteins

can be identified by peptide sequencing using the tandem mass spectrometry (MS/MS) mode of operation in which a mass spectrometer uses two or more mass analyzers.

precursor ions

After an initial mass analysis, individual peptide ions in the mass spectrum can be selected. The selected ions are known as ________.

product ions

A precursor ion is then subjected to the second round of fragmentation through collision and is broken into smaller ions known as _______.

top-down strategy

In ________, intact proteins in a complex mixture are fractionated and separated into less complex protein mixtures or single proteins.

bottom-up strategy

The ________ is commonly used for high-throughput analysis of small peptides derived from highly complex samples.

Sort-then-break approach

Proteins in a complex mixture are first isolated and then enzymatically cleaved.

Break-then-sort approach

Also known as the shotgun approach, wherein proteins are cleaved first and the resulting peptide mixture is then separated by LC and analyzed by tandem MS.

Human Microbiota

Microbial community.

Metagenomics

Allows the study of the microbial genome of complete microbial communities harvested directly from natural environments.

5S and 23S rRNA genes

encode the RNA components of the large subunit of the ribosome.

16S rRNA gene

encodes the RNA component of the small subunit of the ribosome.

intergenic spacer region (ISR)

The _______ between the 16S rRNA and 23S rRNA genes in the rRNA operon is another commonly used marker.

Fluorescence

The emission of light by a fluorophore.

Fluorophore

A moiety in a molecule that fluoresces on absorbing the energy from an excitation light source or radiation.

Raman spectroscopy

Utilizes a near-infrared excitation light source and measures the scattering of laser light caused by the vibrating molecules of a sample.