Crime Scene Unit Presumptive Testing Procedures. Evidence Collection Division

Transcription

1 Crime Scene Unit Presumptive Testing Procedures Evidence Collection Division

2 PRESUMPTIVE TESTING Various chemical processes and presumptive tests are utilized in crime scene investigations to assist in the detection and development of potential evidence. These presumptive tests are useful in determining the possible presence of biological material, but may produce a false positive reaction in the presence of various resembling substances. Should a positive reaction take place, the result only suggests the presence of biological material and that it may be worthy of collection. Preparation of Reagents and Quality Control Checks All reagents will be quality tested to ensure reliability, and all associated lot numbers and QC checks will be documented in the case record. Sampling/Sample Selection The area to be sampled is based on the training and experience of the analyst. Phenolphthalein Test Catalytic tests for blood are based on the peroxidase-like activity exhibited by the heme group of hemoglobin. The test is exceedingly sensitive to minute traces of hemoglobin and its derivatives but can produce a false positive reaction in the presence of any of a number of oxidizing substances. Should a color reaction take place, the result only indicates the possible presence of blood; the test is therefore a presumptive test. Safety Body fluids and extracts may contain infectious agents; therefore universal precautions should be employed during biological evidence handling. Gloves must be worn during testing. This reagent presents the following hazards: o Phenolphthalein solution B: Flammable. Handle with care. Harmful or fatal if swallowed, inhaled, or absorbed through skin. Avoid contact with eyes, skin, and clothing. Light sensitive! Avoid light, heat, sparks, and flame. Keep container tightly closed. o 3% Hydrogen Peroxide Solution C: Handle with care. May be harmful if swallowed, inhaled, or absorbed through skin. Avoid eye contact (irritant). Avoid contact with eyes, skin, and clothing. Equipment, Materials, and Reagents Sterile cotton swab(s) Filter Paper Disposable gloves Phenolphthalein Forensic Test reagents o Solution A: Distilled Water o Solution B: Phenolphthalein o Solution C: 3% Hydrogen Peroxide Controls: o Positive control: Known blood (Blood Stain card) o Negative control: Water Uncontrolled When Printed Page 2 of 5

3 Quality Control Before using phenolphthalein in the field, a positive and negative control must be used to test the reagent. The positive control test will be conducted by transferring a portion of the known blood sample onto a cotton swab and administering the phenolphthalein reagents. A successful positive control is a pink reaction to the known blood standard. A negative control test will be conducted by moistening an unused cotton tip applicator with distilled water. A negative control test should not show any color reaction. This quality control check will be documented in the case record. Record the phenolphthalein, distilled water, and 3% hydrogen peroxide lot numbers in the case record. Procedure 1. Moisten a sterile cotton swab or filter paper with sterile distilled water (Solution A). There is no need to flood the swab or filter paper with water, just use a small amount. Take care not to touch the swab or filter paper to the bottle. 2. Rub or press down on the suspected bloodstain with a sterile cotton swab or filter paper to allow for any possible blood to transfer. 3. Apply approximately 1-2 drops of Phenolphthalein Solution B to the swab or filter paper. 4. Observe briefly to identify a color change. A green color is often observed when blood is present. 5. Add approximately 1 drop of 3% hydrogen peroxide Solution C to the swab or filter paper. 6. Observe the swab or filter paper for an immediate pink color change indicating a positive reaction. 7. After interpretation, record the results in the case record. This is a PRESUMPTIVE test only! If the test is positive for blood, the stain or a sample may be collected for analysis by the Houston Forensic Science Center. If the test is not positive for blood, then a sample of the stain does not need to be collected. Technical Notes If the stain is extremely small or is presumed to be blood by its appearance, the stain may be collected without performing a phenolphthalein test. If no potential blood stains are observed visually, a general swabbing of the area or item can be performed and the phenolphthalein test run on those swabs. Do not perform a general swabbing on areas to be printed. Storage, Labeling, and Expiration The phenolphthalein test kits can be stored at room temperature. Each investigator should have an aliquot of Solutions A, B and C that will be placed in dispenser bottles stored at room temperature. Each bottle will be labeled with appropriate reagent name, lot numbers, and expiration dates. The kits may be used on evidence samples up to the expiration date stated on the bottles. Disposal Dispose of unused mix in a sink under running water. Uncontrolled When Printed Page 3 of 5

4 Leucocrystal Violet Leucocrystal violet (LCV) reacts with the heme or iron group found in blood and gives a blue/purple color reaction. This test can be used to identify bloodstains that are not visible and to enhance the contrast of bloodstain patterns that are visible. Should a color reaction take place, the result only indicates the possible presence of blood; the test is therefore a presumptive test. Safety Equipment, Materials, and Reagents 5-Sulphosalicylic Acid (SSA) 3% Hydrogen Peroxide Sodium Acetate (NaAc) Leucocrystal Violet (LCV) Spray bottle(s) Disposable gloves Positive control: Known blood Negative control: Blank swab or other appropriate substrate Reagent Preparation Add and dissolve 10 grams of 5-sulfosalicylic acid in 500 ml of 3 % hydrogen peroxide. Mix until all powder is completely dissolved. Add 4.4 grams of sodium acetate to the solution. Add 1.1 grams of leucocrystal violet to the solution. Mix until all powder is completely dissolved. Reagents can be adjusted to the volume needed. Document the 5-sulfosalicylic acid, sodium acetate, and leucocrystal violet Lot # s in the case record. Quality Control Prior to use, the LCV solution must be subjected to the appropriate quality control test as outlined below: Spray the LCV solution onto a known blood sample. Spray the LCV solution onto a known non-blood sample (cotton swab or appropriate substrate). The known blood sample must give a blue/purple color reaction indicating a positive result. The cotton swab or other appropriate substrate must not produce a blue/purple color reaction which indicates a negative result. The results of each control test must be recorded on the case notes. Procedure 1. The area of interest should be photographed both before and after the application of LCV solution. 2. Complete the appropriate quality control reagent checks. 3. Lightly mist an area of interest until the first sign of blue/purple color is noted. 4. Development is typically within 30 seconds. Storage, Labeling, and Expiration Do not use leucocrystal violet crystals that are yellow instead of white; this means the crystals are old. The leucocrystal violet, 5-sulfosalicylic acid, and sodium acetate should be stored in a cool and dry area away from incompatible substances. Uncontrolled When Printed Page 4 of 5

5 Premeasured bottles containing LCV reagents should be labeled with the following information: Chemical s name Stock bottle lot number Amount weighed out Date that the LCV reagent(s) was aliquoted Preparer s initials. If stored, the prepared LCV solution has an expiration of 3 months from the date it was mixed together, if stored in a cool and dry environment. If stored, the LCV solution label must include at a minimum: Reagents name Lot number which contains the date prepared and the initials of the preparer (e.g. mmddyyinitials) Expiration date Disposal Dispose of unused mix in a sink under running water. References Lytle, L. T Chemiluminescence in the visualization of forensic bloodstains. Journal of Forensic Sciences 23(3): Saferstein, R. Forensic Science Handbook: Identification and Grouping of Bloodstains, (Prentice-Hall, Inc., 1982), p Cox M A Study of the sensitivity and specificity of four presumptive tests for blood. Journal of Forensic Sciences. 36(5): Kirk P Crime Investigation. 2nd ed. Malabar (FL): Robert E. Krieger Publishing Company. Chapter 15, Blood: general testing; p Uncontrolled When Printed Page 5 of 5