Examination of Bloodstain PDF

# Examination of Bloodstain ## General Questions Five specific questions as guidelines for determining the nature of a crime: 1. Is the sample, blood? 2. Is the sample, animal blood? 3. If the sample is animal blood, from which species did it come from? 4. If the sample is human blood, what type...

# Examination of Bloodstain ## General Questions Five specific questions as guidelines for determining the nature of a crime: 1. Is the sample, blood? 2. Is the sample, animal blood? 3. If the sample is animal blood, from which species did it come from? 4. If the sample is human blood, what type is it? 5. Can the sex, age, and race of the source of blood be determined? ## Physical Examination of Bloodstain ### 1-Color * Differs according to its age and source * Fresh stain ...... bright red * 1-2 days...... reddish brown (old stain) * 2-3 days...... dull brown (older stain) * Blood on leather is brown at once due to formation of met Hb * Blood on glass remain red for several months. ### 2- Clotting * Usually occurs 2-10 minutes after blood shedding due to transformation of soluble fibrinogen into insoluble fibrin. * Blood from dead body is partially coagulate as its plasma has no more fibrinogen, being all slowly changed into fibrin. * It may be delayed in some cases as hemophilia, warfarin toxicity and asphyxia. ### 3- Dryness * Depends on atmospheric conditions * Recent and fresh stain is wet and need 20-30 min to be dried in Egypt. ### 4- Solubility * Fresh stain contains oxy Hb which dissolved in dist. water or normal saline. * Old stain contains met Hb or hematin which dissolved in diluted acid or alkali. * Older stain contains hematoporphyrin which dissolved in strong acid or alkali. * Blood in case of burns need HCL 5% for dissolving. ### 5- Shape * Arterial blood stain is in the form of jets or sprays. * Circular in shape if the blood dropped vertically on the ground. * Pear shape bloodstain indicates wounded moving person, and the pointed end indicate the direction of the movement ## Chemical Examination of Bloodstain ### 1- Preliminary (macrochemical) tests: * **Good negative tests** (if the result is +ve, not confirm that it is blood but -ve result is a definite proof that it is not blood) * They depend on the presence of protein and oxidase enzyme in the blood. * The stain is prepared in the form of clear solution (filtered if turbid). **A- Tests depend on the presence of protein include protein, ammonia and foam (Ganttner's) tests.** ### B- Tests depend on presence of oxidase enzyme * Which can oxidize any oxidizable reagent in the presence of oxygen. * This oxidizable reagents changed in color when oxidized. * Oxidases are also found in other vegetable stains, thus these tests like others, only good negative tests. **These tests include:** Guaiacum, Benzedine, Phenol phethalein (Kastle-Mayers), O-Tolidine and Leucomalachite green tests. ### 2- Confirmatory chemical tests: **Micro-chemical tests:** These tests depend on the presence of hemoglobin in the dried blood and include **Teischmann's (Haemin crystal)** and **Takayam's (Haemochromogen crystal) tests.** #### Teischmann's test (Haemin crystal) * Depend on the presence of Hb in the dry satin. * Hb + Acetic acid → acid hematin * acid hematin + NaCl → haemin crystals * Reagent NaCl 1gm NaBr 1gm Na[e 183 1gm Acetyc acid 100cc **Technique:** Dry stain on clean slide + one drop of reagent, heating gently till appearance of brown color then cooling and examination under microscope. **Haemin crystals:** * Color brown * Shape rhombic * Reaction acidic * Arrangement single or in groups forming Chinese letters appearance. #### Takayama's(Haemochromogen crystal) * Reagent Na OH 1 part Pyridine 1 part Saturated solution of glucose 1 part Distilled water 7 parts **Technique:** Dry stain on clean slide + one drop of reagent, heating gently till appearance of pink color then cooling and examination under microscope. **Haemochromogen crystals:** * Color pink * Shape spindle * Reaction alkaline * Arrangement in groups forming feather or rosette like appearance. ## Microscopical examination of bloodstain * It considered as confirmatory tests of bloodstain. * Depend on the presence of blood corpuscles. * Used in case of fresh blood blood film is made and stained with Leishman or Giemsa stain. **The prepared blood film can determine the followings:** ### 1- Species from which the stain is derived: * **Mammalian RBCs** (except camel) are circular, biconcave and non nucleated discs. * **Camel RBCs** are oval biconvex and non nucleated. * **Non mammalian RBCs** (as frog, birds and reptiles) are oval, biconvex and nucleated. ### 2- Type of animal by measuring RBCs diameter: * Sheep and goat → 5.5 micron * Horse, cat, ox and pig → 6 micron * Human, rabbit and dog → 6.5 micron * Elephant → 9.5 micron ### 3- Sex 3% of female neutrophil contain Davidson body (drumstick consists of a small nuclear mass, about 1.5µ in diameter which is attached to the body of the nucleus by means of a thin filament). ### 4- Toxicity In case of lead toxicity, pyremidine -5- nucletidase enzyme which responsible for removal of nuclear remnant of developing RBCs in bone marrow is inhibited by lead. In this case, the formed RBCs contain basophilic fragments which appear in blood film as punctuating or stippling RBCs. ## Spectroscopic examination of bloodstain one of the confirmatory, simplest tests. ### Advantages of Spectroscopic examination * It can be done on a very small quantities of fresh or dry blood * The sample can be used in other chemical tests. * This test can be used in the diagnosis of some cases of toxicity as CO, nitrite, cyanide and H₂S in addition to determination of stain age. ### Principle of Spectroscopic examination * Hemoglobin and its derivatives have characteristic absorption bands in the spectrum. * Other dyes may give absorption bands similar to Hb or one of its derivatives. * Addition of any oxidizing or reducing agent do not altered the bands of the dye while altered the bands of Hb due to its change by the oxidizing or reducing agent. ### Method. adjust the apparatus till the Fraunhaufer's lines are quite distinct and sharp. The stain fluid is interposed between the apparatus and the source of white light. ### Result: * Oxy Hb shows 2 dark bands between D and E lines ### Carboxy Hb. In case of carbon monoxide toxicity, its bands similar to those of oxy Hb but with slight shifting to right side. To differentiate between them add reducing agent which change oxy Hb into reduced one while carboxy Hb remain unchanged. ### Reduced Hb. obtained by lifting the stain for an hour in warm weather or 2 hours in cold one, the blood acquires purple color where Oxy Hb changed to reduced Hb. Only one broad badly defined band absorbing most of the spectrum between D and E lines will appear. ### Met. Hb. obtained by addition of few drops of pot. Ferro cyanide, pot. Permanganate or some other oxidizing agents. Four absorption bands, one close to D line, the second terminate slightly behind E line, the third band lies between C, D lines, the fourth between E and F lines. ### Hemochromogen Two distinct absorption bands, one at the midline between D, E lines while the other band lies on the left of E line. ## Serological examination of bloodstain ### 1- Precipitin test: This test is based on the fact that injection of certain protein (serum) into living animal causes the formation of antibodies in the blood of this animal, these antibodies have a highly specific action when come in contact with the substance with which the animal injected. ### **SEROLOGICAL EXAMINATION** ### 1. Precipitin Test: The antibodies causing this reaction are known as precipitins, and the animal serum is known as an anti-human precipitin serum. A rabbit or a fowl is injected with human blood every third day for 3-5 injections. After this the animal is killed, and the antiserum is collected. A suitable antiserum should react immediately or within a minute on the 1:1,000 dilution. ### 2- Blood grouping With the help of **blood grouping**, we can say that a certain blood stain did not come from a certain individual!! * **Group A** * **Group B** * **Group AB** * **Group O** - **ABO blood group antigens present on red blood cells** - **IgM antibodies present in the serum.** ### Blood Typing Blood typing requires 2 antiserums: 1. anti-A 2. anti-B. Insert a droplet of these antiserums in samples of blood can determine which samples maintain normal appearance and which become clotted, or agglutinated under microscopic examination. * 'A' blood agglutinated by anti-A serum. * 'B' blood agglutinated by anti-B serum. * 'AB' blood by both. * 'O' blood by neither. ### • DNA fingerprinting examination