Term
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Definition
8% of total body weight
5-8 pints for average adults
Loss of greater than or equal to 40% of blood volume is fatal |
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Term
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Definition
Plasma
Can clot
Serum is the remaining fluid after the clot is removed |
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Term
| Cellular portion (suspeded in the plasma) |
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Definition
Red blood cells - erythrocytes, no DNA, hemoglobin
White blood cells - leukocytes, immunity, DNA
Platelets - thrombocytes, blood clotting, no DNA |
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Term
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Definition
Adult Hb is a heterodimer
2 alpha and 2 beta chains
Heme moiety (ferroprotoporphyrin binds O2) |
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Term
| Heme moiety of a RBC Hemoglobin |
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Definition
Composed of protoporphyrin (X and ferrous Fe2+ iron)
Fe 2+ binds to protoporphyrin, the hemoglobin chain, a molecule of O2, and CO & Cyaninde (leads to asphyxia) |
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Term
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Definition
Preliminary / Presumptive Assays
Confirmatory Assays |
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Term
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Definition
Sensitive, quick, and easy
Screening tool
Not specific
Based on redox reactions catalyzed by the heme moiety
Results in color change or release of a photon by chemiluminescence or fluorescence |
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Term
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Definition
Performed by a lab
Greater specificity than presumptive assays - for determing animal v human
Based on crystal formation, primary serological reactions, spectrophotometry, or RNA-based assays
Rarely performed because we jump to DNA |
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Term
| Presumptive Assays: Colorimateric Assays |
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Definition
Pheonlphthalein: Kastle-Meyer Reagent
Leucomalachite Green (LMG)
Ortho-tolidine
Tetramethylbenzidine (TMB) |
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Term
| Method of Presumptive Assays |
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Definition
Sample stain with clean, moistened cotton swab
Add a drop of reagent
Add a drop of H2O2
Watch for color development
Heme is ALWAYS the catalyst |
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Term
| Presumptive Assay: Kastle-Meyer |
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Definition
Phenolphthalin = reduced, colorless
Phenolphthalein = oxidized, pink |
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Term
| Presumptive Assay: Leucomalachite Green (LMG) |
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Definition
Leucomalachite green = reduced, colorless
Malachite green = oxidized, blue-green |
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Term
| Presumptive Assay: Othro-Tolidine |
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Definition
Ortho-Tolidine = reduced, colorless
O-Tolidine = oxidized, blue
Replaced by TMB
Produces a false positive with some gun metals |
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Term
| Presumptive Assay: Tetramethylbenzidine (TMB) |
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Definition
Colorless as reduced
Blue-green as oxidized
Hemastix: reduces ability to recover DNA, gives false positive with some gun powder residues |
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Term
| Chemiluminescent Presumptive Assays |
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Definition
Product of the chemical reaction is the emisson of light
Luminol, Bluestar |
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Term
| Chemiluminescent Presumptive Assay: Luminol |
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Definition
Emits blue color: detects cleaned up / patterns of / small traces of blood
Performed in darkness
May dilute sample
False positives: bleach, plant peroxidase (ie: horseraddish), copper, copper-containing alloys, iron peroxidases, cyanide, feces, blood-containing urine
Only lasts a few hours after it is mixed |
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Term
| Chemiluminescent Presumptive Assay: Bluestar |
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Definition
Luminol-based
Reveals blood that is washed out, wiped off, or invisible to the naked eye
Does NOT alter DNA, safer & easier to prepare, comes in a dissolveable tablet (longer shelf life)
Same false positives as luminol |
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Term
| Fluroescence Presumptive Assays |
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Definition
Absorption of UV or visible radiation puts electrons into a higher orbital
When electrons drop down to original ground states, energy is released (transferred to vibrational & rotational energy of molecular bonds) or energy is released as a photon f lower energy wavelength (fluorescence)
Fluorescein |
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Term
| Fluroescence Presumptive Assays: Fluoresceine |
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Definition
Oxidized by the peroxidase acivity of heme in the presence of H2O2 = fluoresce under blue-purple light at 425, 485 nm wavelengt of light from ALS
Emits a ellow-green coor: used in opthamology; topical fluoresceine to diagnose corneal abrasions, ulcers, infections |
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Term
| Factors Affecting Presumptive Assay Results |
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Definition
Oxidizing agents - false positives; metal salts, bleach, household cleaners
Plant peroxidases - false positives; most are heat labile, but heme is not heat labile
Reductants - false negative |
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Term
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Definition
Microcrystal Assays
Chromatographic & electrophoretic methods
RNA-based methods
Spectrophotometric Methdos
Immunological methods |
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Term
| Confirmatory Assays: Microcrystal Assays -> Hemochromagen Crystal Assay (TAKAYAMA) |
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Definition
Method: small amount of putative blood added to a slide
Add pyridine and glucose
Under alkaline conditions
Slide heated to form pyridine ferroprotoporphyrin crystals in the presence of blood
View under a microscope |
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Term
| Confirmatory Assays: Microcrystal Assays -> Hematin Crystal Assay (TEICHMAN) |
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Definition
Small amount of putative blood added to a slide
Add glacial acetic acid and salts
Slide heated to form ferriprotoporphyrin chlordie (hematin chloride) crystals in the presence of blood
Viewed under the microscope |
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Term
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Definition
Screening
Past: serological techniques
Today: DNA quantitation
Can be used at the crime scene to eliminate non-human samples
Necessary in animal cases
Use primary and secondary binding assays |
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Term
| Species Identification: Types of Antibodies |
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Definition
Antihuman serum is polyclonal
Albumin is the most abundant serum protein
Hemoglobin antibody
Glycophorin A (GPA) -> RBC membrane of Ag |
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Term
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Definition
Series of dilutions of antiserum are made and each dilution is then tested for activity using either precipitation or agglutination methods
Titer
Serial dilution |
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Term
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Definition
The reciprocal of the highest dilution giving a positive reaction
Dependent upon the animal because the known value being compared is dependent upon the animal
In pharmeceuticals, titer is a cut-off. Higher titer = still protected, lower titer = not protected |
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Term
| Specificity of Anti-human Sera |
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Definition
Usually cross-reacts with higher primates
Must be validated for cross-reactivity and tissue specificity
Reactive with other human fluids, such as semen and saliva |
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Term
| Optimal Conditions for Testing |
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Definition
| Proper buffer system, ionic strength, intro of polymers, temperature, pH, and sample preparation (extracted with buffer or saline, controls). |
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Term
| Immunochromatographic Assays for Blood |
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Definition
Identification of hemoglobin protein
-Hexagon OBTI
- ABAcard HemaTrace
Identification of human glycophorin A protein
- RSID-blood |
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Term
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Definition
Immunochromatographic Assay
Precipitation-based Assays
Ring Assays
Ouchterlony
Crossed-over Electrophoresis |
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Term
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Definition
| Identity, nonidentiy, partial identity |
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Term
| Techniques for Blood Group Typing: Agglutination Assays |
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Definition
Lattes Crust Assay
Absorption-Elution Assay
Developed due to limitations with blood group typing
Inherited protein polymorphic markers (amino acid sequence of proteins vary in the population)
Identification: electrophoretic separation based on MW & change of protein variants |
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Term
| How many proteins in humans are polymorphic? |
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Definition
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Term
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Definition
Agglutination Assay
Simple & rapid - but NOT sensitive
Requires a lot of blood
NOT reliable for old stains |
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Term
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Definition
Highly sensitive
Can be used on old stains
Ag immobilized on solid matrix phase. Ab added and absorbed, unbound is washed away. Bound Ab is eluted and tested with indicator cells & identified by a positive agglutination reaction. |
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Term
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Definition
Transports oxygen
Useful for markers |
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Term
| How many variants of hemoglobin have been identified? |
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Definition
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Term
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Definition
Two alpha and two gamma chains
Replaced after birth to be two alpha and two beta chains
Can determine infanticide based on hemoglobin |
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Term
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Definition
Sickle Cell
Investigative leads for indication of ethnic origin |
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Term
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Definition
Forms stable complexes with hemoglobin
Controls hemoglobin excretion fom the body via the liver |
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Term
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Definition
Multiple molecular forms
Ex: phosphoglucomatase (PGM), peptidase A, transferrin, and 11+ more |
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Term
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Definition
| Occurrence in a population of two or more genetically determined alternative phenotypes with frequencies greater than could be accounted for by mutational drift |
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Term
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Definition
A group of antigens produced by allelic genes at a single locus and inherited independently of any other genes.
Antigen polymorphisms that are present on the erythrocyte surface. |
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Term
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Definition
A, AB, B, O
Type A: A antigen
Type B: B antigen
Can be found in saliva and semen and other tissues
Can exclude suspects, but cannot ID them |
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Term
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Definition
Almost everyone produces it
Fucosyltransferase |
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Term
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Definition
Generated by some
Galactosyltransferase |
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Term
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Definition
Encoded by human FUT1 and FUT2 loci
Adds fucose to the end of a glycolipid or glycoprotein |
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Term
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Definition
| Glycolipid on erythrocytes |
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Term
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Definition
| Glycoprotein on other tissues & in secretions |
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Term
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Definition
Adds H antigen
O allele = null / non-functional
A allele = A-transferase (adds N-acetylgalactosamine)
B allele = B-transferase (adds galactose) |
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Term
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Definition
A, B, O antigens widely distributed in other tissues
Secretion under control of FUT2 gene - if functional, H antigen is distrubuted on surface of cells of tissues other than blood & ABO antigens can be added onto these tissues
75% of Caucasions |
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Term
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Definition
Homozygous for a mutation in FUT2 resulting in a truncated protein
20% of Caucasions
FUT2 no longer works |
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Term
| Forensic Protein Profiling |
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Definition
Developed due to limitations with blood group typing
Inherited protein polymorphic markesr because amino acid sequence of proteins varies in the population |
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