1. Presumptive blood tests

2. Presumptive vs confirmatory blood tests
Presumptive Tests (Also known as preliminary tests, screening tests or field tests)
Indicates a substance is present
Not specific
Provide initial information to determine what test to perform next
used in combination with confirmatory tests
Confirmatory Tests
Confirm a substance is present
specific
May be more expensive, require additional equipment, and take longer

3. tests Kastle‐Meyer (Phenolphthalein) Leucomalachite Green (LMG)
Hemastix®
Tetramethylbenzidine (TMB)
Luminol
Fluorescein

4. Swab the stain using a slightly moistened (with distilled water) swab
Kastle‐Meyer Test
The Kastle-Meyer solution is a phenolphthalein solution which has been reduced, usually by reacting it with powdered zinc. The basis of the test is that the heme in blood reacts with the colorless phenolphthalein into bright pink phenolphthalein.
Place a small cutting, swabbing, or extract of the suspected bloodstain on filter paper
‐OR ‐
Swab the stain using a slightly moistened (with distilled water) swab
Add two drops of reagent and wait for about five seconds.
Add two to three drops of 3% H2O2.
If immediate color change to PINK–the test is POSITIVE for the possible presence of blood
If no color change –blood is not present or is in too limited quantity for the test to detect.
Note: The swab will eventually turn pink (even if negative) over time due to nature of oxidation reactions.

5. Kastle-Meyer limitations
Sensitivity: 1:1000 on dried stains
Specificity
Can weed out false positives between steps 2 and 3
False positive in presence of cauliflower or broccoli
Will not detect differences in animal or human blood
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6. Leucomalachite Green –How to Perform
Hemoglobin catalyzes the reaction between LMG and hydrogen peroxide, converting the colorless LMG into malachite green. Therefore, the appearance of a green color indicates the presence of blood
Place a small cutting, swabbing, or extract of the suspected bloodstain on filter paper
‐OR ‐
Swab the stain using a slightly moistened swab
Add one to two drops of LMG reagent.
Note color change: If there is a color change, the test is considered inconclusive.
2. Add one to two drops of H2O2.
Note the results.
If color change to deep green‐blue, the test is positive for the possible presence of blood.
If no color change the test is negative.

7. Leucomalachite Green Limitations
Sensitivity: 1:1000
Specificity: Chemical oxidants, vegetable peroxidases
Will not detect differences in animal or human blood

8. Hemastix® Reagent strips React with hemoglobin in blood
Reagent on Hemastix® is diisopropylbenzene dihydroperoxide and 3,3’,5,5’‐tetramethylbenzidine (TMB)
•Color change ranges from orange to green to Possibly blue with higher concentrations of blood
1. Slightly moisten the pad on the tip of the strip with water.
2. Rub the damp pad on the stain in question.
Note any color change within 60 seconds and compare to the chart on the bottle. More green indicates more hemoglobin

9. Hemastix limitations Sensitivity = 0.015 ‐0.062 mg/dL free hemoglobin
Specificity: Chemical oxidants, vegetable peroxidases
Will not detect differences in animal or human blood c

10. Tetramethylbenzidine (TMB)
Reacts with heme to produce a color change
Place a small cutting, swabbing, or extract of the suspected bloodstain on filter paper
‐OR ‐
Swab the stain using a slightly moistened swab
Add one drop of TMB solution.
Add one drop of 3% H2O2.
Detect color change:
If the stain turns blue‐ish green, the test result is POSITIVE for the possible presence of blood
NEGATIVE if no color change

11. TMB Limitations Sensitivity1:10,000 on dried stains
Specificity: Not as specific as Kastle-Meyer test
False positives to vegetable peroxidases, bleach, potassium permanganate
Will not detect differences in animal or human blood

12. Luminol
The iron from the hemoglobin in the blood starts the chemiluminescence reaction that causes luminol to glow, so a blue glow is produced when the solution is sprayed where there is blood. Only a tiny amount of iron is required to catalyze the reaction.
Spray the luminol directly onto the stain in question. If the stain emits a light then the test result is POSITIVE for the possible presence of blood
If there is no reaction the result is NEGATIVE
Note: This test needs to be done in the dark to see the luminescence reaction, which can last for approximately 15 seconds.

13. Luminol limitations Sensitivity: (10-6 to 10-8 )
Specificity: Many false positives –bleach, metals, chemical oxidants, vegetable peroxidases
Will not detect differences in animal or human blood
•Mostly used at crime scene
Can dilute out stain (possibly too much for DNA analysis)
Used more for blood spatter, crime scene reconstruction

14. Fluorescein
Reacts with red blood cells to glow
Needs an alternative light source to visualize
Reveals trace amounts of blood
Spray the fluorescein directly onto the stain in question. If the stain emits a light then the test result is POSITIVE for the possible presence of blood
If there is no reaction the result is NEGATIVE
Note: This test needs to be done in the dark to see the luminescence reaction, which can last for approximately 5-7 minutes.