CART

Staining blood traces

Staining solutions and reagents used for improving the contrast of marks (fingerprints and footmarks) made in blood.
 

Acid Yellow 7

Main uses: Blood traces on non-porous surfaces
 
Acid Yellow 7 is a dye solution in a water/acetic acid/ethanol mixture that is used for staining fingerprints and shoeprints made in blood. Prints in blood are colored yellow after treatment with Acid Yellow 7. They then fluoresce under blue/blue-green light. Acid Yellow 7 should not be used on absorbent surfaces like paper, carton material, bed sheets, or carpet. It works very well on non-absorbent backgrounds like linoleum, glass, tiles, painted surfaces, or PVC floor covering.
Product #
Description
Quantity
Add
 
 
B-88400
Acid Yellow 7, 500 ml staining solution (water/ethanol/acetic acid based), in spray bottle
 
B-89825
Acid yellow 7, 25 g
 
Blood fixative
B-89700
Fixative for traces in blood (2% 5-sulphosalicylic acid in water). In spray bottle, 500 ml
 
B-89701
Fixative for traces in blood (2% 5-sulphosalicylic acid in water). In polyethylene bottle, 100 ml
 

On what surfaces can it be used

Acid Yellow 7 is a dye solution in a water/acetic acid/ethanol mixture that is used for staining fingerprints and shoeprints made in blood [1]. Prints in blood are colored yellow after treatment with Acid Yellow 7. They then fluoresce under blue/blue-green light. Acid Yellow 7 should not be used on absorbent surfaces like paper, carton material, bed sheets, or carpet. It works very well on non-absorbent backgrounds like linoleum, glass, tiles, painted surfaces, or PVC floor covering.

Before staining, prints in blood should be fixed to prevent them from running (causing loss of detail) when the staining solution is applied. In general fixative is applied before any staining solution except Leuco Crystal Violet, which contains fixative.

How to fix the prints

Fixing is best done with a 2% solution of sulfosalicylic acid in water (20-23 gram in 1 liter). To make sure the print in blood is thoroughly fixed, we recommended the following method using absorbent paper (like filter paper, tissue paper, or paper towels) and a wash bottle containing sulfosalicylic acid solution.

Take a dry piece of the absorbent paper that is sized to cover the print(s). Hold it directly over the print-area, parallel to the surface and just slightly above the surface. To begin fixing, drop one edge of the paper to the surface and moisten heavily along its entire length, so that it is anchored and will stay in place.

Starting from that edge, wet the paper progressively further while smoothing the wetted part onto the print and minimizing trapped air bubbles. Work carefully from one edge to the other. When possible, for example, with an object or with a print on an angled surface start from the top and work downwards.

Once the wet paper entirely covers the print, leave it there for a minimum of three minutes.When the blood is a thick layer, leave the paper there 5 minutes or more. Remove the paper. An excess of fixative can be rinsed away with water, but this is not necessary.
When the blood is relatively fresh, you will notice that fixing changes its color from dark red to dark brown.

Staining procedure

Once the print is fixed, it can be stained with Acid Yellow 7 solution. Use a wash bottle to apply the solution to the article or area of interest. With fingerprints, a disposable pipet can be used. Alternatively you can submerge the article in a container/bath of solution if the size of the article permits.

Spraying is technically feasible but will likely raise the amount of ethanol vapor in the air. Sufficient protection measures should be taken to avoid inhalation of aerosols.
After the staining solution is applied leave it in contact with the print for 1-3 minutes. Then wash the surface with the same water/acetic acid/ethanol mixture as used for the dye solution, but without the dye.

Alternatively, water can be used but may result in more background staining. If the print is located on a floor, the wash solution might be removed with a vacuum cleaner that can handle water. Otherwise, remove it with paper towels or the like.

Lifting and fluorescence

Acid Yellow 7 not only has a good staining capacity but has some special characteristics.The stained prints will fluoresce when excited with blue and blue-green light though the fluorescence will be faint. To obtain the best result, eliminating backgrounds and/or background fluorescence, stained prints can be lifted with a white BVDA Gellifter.

Before lifting with the white Gellifter, the area should be completely dry. Be very careful not to trap air bubbles under the lifter surface. Leave the lifter on the print for only a short time, something like a minute. Photograph within a couple of hours after lifting as the dye will slowly diffuse in and across the surface of the lifter, thereby blurring the print.
Lifting can be done more than once, with or without re-staining between lifts. Repeat lifting/staining in important cases.

Fluorescence is excited with blue and blue-green light (400-490 nm) and visible even when using long-wave UV. The filter used in front of the lens depends on the excitation wavelength: clear for UV to yellow/light orange for the higher wavelengths.

Safety

Acid Yellow 7 powder is not labeled as a hazardous preparation. Since its working solution is based on a water/ethanol mixture, no harmful fumes are liberated in normal use, although the acetic acid smell might be irritating. When large amounts are used and/or the temperature is high enough, a potentially explosive air/ethanol vapor mixture can result unless ample ventilation is provided. Acid Yellow 7 solution will color the skin and clothing.

Sulfosalicylic acid crystals are labeled as an irritant substance but the 2% solution is not labeled as a hazardous product. Given the fixative effect on blood, we strongly advise wearing gloves (vinyl gloves will suffice) when using it.

Avoid breathing aerosols (mists) and vapors of the fixative, washing liquid, and Acid Yellow 7 solution. Likewise avoid contact with solutions and the Acid Yellow 7 powder. Wash your hands after each session, especially before eating or drinking.

Composition of the solutions


Acid Yellow 7

1 gram Acid Yellow 7
50 mlacetic acid (glacial, 99%)
250 mlethanol (98% or higher)
700 mldemineralized or distilled water
--------
1000 ml working solution

Preferably use an Erlenmeyer flask for preparation of the Acid Yellow 7 solution. Add water first and dissolve the Acid Yellow 7 powder in the water by swirling the flask or using a magnetic stirrer and a PTFE-covered stir bar. The powder will dissolve quickly. Then add ethanol and acetic acid (order not important.)


Blood fixative

20 grams 5-Sulfosalicylic acid, dihydrate 
1000 mldemineralized or distilled water 
-------- 
1000 ml working solution

Add components to a beaker/flask of suffient size and mix till complete dissolution, using a magnetic stirrer.


Washing solution

50 mlacetic acid (glacial, 99%)
250 mlethanol (98% or higher)
700 mldemineralized or distilled water
--------
1000 ml  washing solution

References and footnotes

[1] Sears, V.G.; Butcher, C.P.G.; Fitzgerald, L.A. "Enhancement of Fingerprints in Blood, Part 3: Reactive Techniques, Acid Yellow 7, and Process Sequences." Journal of Forensic Identification 2005, Vol. 55, No. 6, p. 741-763.



Name:
Acid Yellow 7 (CI 56205), 6-Amino-2,3-dihydro-1,3-dioxo-2-(4-methylphenyl)-1H-benz[de]isoquinoline-5-sulfonic acid sodium salt
CAS No.: 2391-30-2
 

Acid violet 17

Main uses: Blood traces on non-porous, semi-porous and porous surfaces.
 

Acid Violet 17 (AV17) is a dye solution based on a water/acetic acid/ethanol mixture that is used for staining fingerprints and shoeprints made in blood. Prints in blood are colored purple after treatment with Acid Violet 17 (similar color as LCV or Coomassie Blue). Acid Violet 17 can be used on absorbent surfaces but will stain the background. The CAST manual suggests to test a small part of the item away from the mark with Amido Black and AV17 to see which one gives less background staining.
Acid Violet 17 can be used after Acid Yellow 7 but may show less contrast than would be obtained with AV17 alone.

Before staining, prints in blood should be fixed to prevent them from running (causing loss of detail) when the staining solution is applied. In general fixative is applied before any staining solution except Leuco Crystal Violet, which contains fixative.

Product #
Description
Quantity
Add
 
 
B-88200
Acid Violet 17, 500 ml in a spray bottle.
 
B-88195
Acid Violet 17, 25 g
 
Blood fixative
B-89700
Fixative for traces in blood (2% 5-sulphosalicylic acid in water). In spray bottle, 500 ml
 
B-89701
Fixative for traces in blood (2% 5-sulphosalicylic acid in water). In polyethylene bottle, 100 ml
 

On what surfaces can it be used

Acid Violet 17 (AV17) is a dye solution based on a water/acetic acid/ethanol mixture that is used for staining fingerprints and shoeprints made in blood [1]. Prints in blood are colored purple after treatment with Acid Violet 17 (similar color as LCV or Coomassie Blue). Acid Violet 17 can be used on absorbent surfaces but will stain the background. The CAST manual suggests to test a small part of the item away from the mark with Amido Black and AV17 to see which one gives less background staining.
Acid Violet 17 can be used after Acid Yellow 7 but may show less contrast than would be obtained with AV17 alone.

Before staining, prints in blood should be fixed to prevent them from running (causing loss of detail) when the staining solution is applied. In general fixative is applied before any staining solution except Leuco Crystal Violet, which contains fixative.

How to fix the prints

Fixing is best done with a 2% solution of sulfosalicylic acid in water (20-23 gram in 1 liter). To make sure the print in blood is thoroughly fixed, we recommended the following method using absorbent paper (like filter paper, tissue paper, or paper towels) and a wash bottle containing sulfosalicylic acid solution.

Take a dry piece of the absorbent paper that is sized to cover the print(s). Hold it directly over the print-area, parallel to the surface and just slightly above the surface. To begin fixing, drop one edge of the paper to the surface and moisten heavily along its entire length, so that it is anchored and will stay in place.

Starting from that edge, wet the paper progressively further while smoothing the wetted part onto the print and minimizing trapped air bubbles. Work carefully from one edge to the other. When possible, for example, with an object or with a print on an angled surface start from the top and work downwards.

Once the wet paper entirely covers the print, leave it there for a minimum of three minutes.When the blood is a thick layer, leave the paper there 5 minutes or more. Remove the paper. An excess of fixative can be rinsed away with water, but this is not necessary.
When the blood is relatively fresh, you will notice that fixing changes its color from dark red to dark brown.

Staining procedure

Once the print is fixed, it can be stained with Acid Violet 17 solution. Use a wash bottle or the spray bottle in which it is supplied to apply the solution to the article or area of interest. With fingerprints, a disposable pipet can be used. Alternatively you can submerge the article in a container/bath of solution if the size of the article permits.

Spraying is technically feasible but will likely raise the amount of ethanol vapor in the air. Sufficient protection measures should be taken to avoid inhalation of aerosols.
After the staining solution is applied leave it in contact with the print for 1-3 minutes. Then wash the surface with the same water/acetic acid/ethanol mixture as used for the dye solution, but without the dye.

Alternatively, water can be used but may result in more background staining. If the print is located on a floor, the wash solution might be removed with a vacuum cleaner that can handle water. Otherwise, remove it with paper towels or the like.

Lifting and fluorescence

Acid Violet 17 can be lifted with a white gelatin lifter (takes much more time than Acid Yellow 7 - around 15 minutes instead of a few minutes with AY7.) The stained prints will fluoresce when excited with green light though the fluorescence will often be faint. To obtain the best result, eliminating backgrounds and/or background fluorescence, stained prints can be lifted with a white BVDA Gellifter.

Before lifting with the white Gellifter, the area should be completely dry. Be very careful not to trap air bubbles under the lifter surface. Leave the lifter on the print for quite some time, something like 15 minutes. Photograph within a couple of hours after lifting as the dye will slowly diffuse in and across the surface of the lifter, thereby blurring the print.
Lifting can be done more than once, with or without re-staining between lifts. Repeat lifting/staining in important cases.

Fluorescence is excited with green light (480-545 nm). A red filter is used in front of the lens.

Safety

Acid Violet 17 powder is not labeled as a hazardous preparation. Since its working solution is based on a water/ethanol mixture, no harmful fumes are liberated in normal use, although the acetic acid smell might be irritating. When large amounts are used and/or the temperature is high enough, a potentially explosive air/ethanol vapor mixture can result unless sufficient ventilation is provided. Acid Violet 17 solution will color the skin and clothing.

Sulfosalicylic acid crystals are labeled as an irritant substance but the 2% solution is not labeled as a hazardous product. Given the fixative effect on blood, we strongly advise wearing gloves (vinyl gloves will suffice) when using it. Wearing protective goggles is not a bad idea either.

Avoid breathing aerosols (mists) and vapors of the fixative, washing liquid, and Acid Violet 17 solution. Likewise avoid contact with solutions and the Acid Violet 17 powder. Wash your hands after each session, especially before eating or drinking.

Composition of the solutions


Acid Violet 17

1 gram Acid Violet 17
50 mlacetic acid (glacial, 99%)
250 mlethanol (98% or higher)
700 mldemineralized or distilled water
--------
1000 ml working solution

Preferably use an Erlenmeyer flask for preparation of the Acid Violet 17 solution. Add water first and dissolve the Acid Violet 17 powder in the water by swirling the flask or using a magnetic stirrer and a PTFE-covered stir bar. The powder will dissolve quickly but due the strong color of the solution this will be hard to see. Allow enough time to dissolve all the dye. Then add ethanol and acetic acid (order not important.) Dissolving the Acid Violet 17 first in the ethanol/acetic acid mixture and then diluting it with water is also possible (as is dissolving it in the complete mixture of course.)


Blood fixative

20-23 grams 5-Sulfosalicylic acid, dihydrate
1000 mldemineralized or distilled water
--------
1000 ml working solution

Add components to a beaker/flask of suffient size and mix till complete dissolution, using a magnetic stirrer.


Washing solution

50 mlacetic acid (glacial, 99%)
250 mlethanol (98% or higher)
700 mldemineralized or distilled water
--------
1000 ml washing solution

References and footnotes

[1] Sears, V.G.; Butcher, C.P.G.; Prizeman, T.M. "Enhancement of Fingerprints in Blood - Part 2: Protein Dyes" Journal of Forensic Identification 2001, Vol. 51, No. 1, p. 28-38.


Name: Acid Violet 17, CI 42650, Coomassie Violet R-150, Benzenemethanaminium, N-[4-[[4-(diethylamino)phenyl][4-[ethyl[(3-sulfophenyl)methyl]amino]phenyl]methylene]-2,5-cyclohexadien-1-ylidene]-N-ethyl-3-sulfo-, inner salt, sodium salt (1:1)
CAS No.: 4129-84-4
 

Amido Black

Amido Black (Acid Black 1) is a dye that stains the protein component of blood blue-black. Amido Black staining solution can be methanol- or water-based. Amido Black in methanol has a greater staining power (compared to the water/citric acid based formulation), but due to the toxicity of the methanol, it is also more dangerous.
 
The formulation from CAST (The Centre for Applied Science and Technology - Home Office - UK) that is based on water/ethanol/acetic acid stains as good as the methanol-based one. For use on a crime scene (for example, shoeprints in blood) the water-based staining solutions are advised.
 
When used on porous or semi-porous surfaces Amido Black will likely produce some background staining. Amido Black will not produce fluorescence.
Product #
Description
Quantity
Add
 
 
B-89500
Amido Black set, water-based. The set consists of two 500 ml spray bottles, one with Amido Black staining solution (B-89600) and the other with a blood fixative (2% 5-sulphosalicylic acid in water, B-89700).
 
B-89501
Amido Black set, methanol-based. Apart from fixative (B-89700) and staining solution (500 ml in spray bottle, B-89601), the set also contains two rinsing solutions for removing background staining: 500 ml methanol/acetic acid and 500 ml 5% aqueous acetic acid.
 
B-88300
Amido Black staining solution, water/ethanol/acetic acid based. This formulation stains as good als the methanol based one (B-89601) but does not have the toxicity problem due to the methanol base. Just as other formulations traces in blood need to be fixed prior to staining (blood fixative:2% 5-sulphosalicylic acid in water, B-89700).
 
B-89400
Amido Black powder, 25 g in a brown glass bottle.
 
Blood fixative
B-89700
Fixative for traces in blood (2% 5-sulphosalicylic acid in water). In spray bottle, 500 ml
 
B-89701
Fixative for traces in blood (2% 5-sulphosalicylic acid in water). In polyethylene bottle, 100 ml
 



Name: Amido Black 10B, Acid Black 1, Naphthol Blue Black, CI 20470, 4-Amino-5-hydroxy-3-(p-nitrophenylazo)-6-(phenylazo)-2,7-naphthalenedisulfonic acid disodium salt
CAS No.: 1064-48-8
 

Crowle's stain

Crowle's Stain, is a protein stain like for example Acid Violet 17 and Amido Black. Crowle's Stain colors marks in blood red. It is also known as Crowle's double stain because it contains two dyes: Crocein Scarlet 7B (major component) and Coomassie Brilliant Blue R (minor component).
 
The formulation contains a toxic substance (trichloroacetic acid) and acetic acid but is otherwise water-based.
 
The stained marks can hardly be lifted with a white gelatin lifter and do not fluoresce, either on the treated surface or the gellifter. In comparisons Crowle's stain gives less contrast than Amido Black or Acid Violet 17.

Product #
Description
Quantity
Add
 
 
B-88500
Crowle's Stain, 500 ml in a spray bottle
 


Name: Crocein scarlet 7B, CI Acid Red 71, 1-Naphthalenesulfonic acid,7-hydroxy-8-[2-[2-methyl-4-[2-(2-methyl-4-sulfophenyl)diazenyl]phenyl]diazenyl]-, sodium salt (1:2)
CAS No.: 6226-76-2
 

Name: Coomassie Brilliant Blue R, CI Acid Blue 83, Benzenemethanaminium, N-(4-((4-((4-ethoxyphenyl)amino)phenyl)(4-(ethyl((3-sulfophenyl)methyl)amino)phenyl)methylene)-2,5-cyclohexadien-1-ylidene)-N-ethyl-3-sulfo-, inner salt, sodium salt (1:1)
CAS No.: 6104-59-2
 

Hungarian Red

Main uses: Blood traces on non-porous surfaces
 
Hungarian Red is a water-based staining solution for traces in blood. It has a number of advantages compared to other staining solutions. It is safe (water-based), stains well, and can be lifted with a white gelatin lifter. A special characteristic of Hungarian Red is that the lifted traces fluoresce under green light, making it possible to visualize weak traces, even when they are present on a dark surface.
 

Product #
Description
Quantity
Add
 
 
B-88000
Hungarian Red set, 100 ml fixative (2% 5-sulphosalicylic acid in water) and 100 ml staining solution, both in 100 ml polyethylene bottles
 
B-88001
Hungarian Red staining solution, 100 ml in a polyethylene bottle
 
B-88100
Hungarian Red set, 500 ml staining solution and 500 ml fixative, both in 500 ml spray bottles
 
B-88101
Hungarian Red staining solution, 500 ml staining solution, in spray bottle
 
Blood fixative
B-89700
Fixative for traces in blood (2% 5-sulphosalicylic acid in water). In spray bottle, 500 ml
 
B-89701
Fixative for traces in blood (2% 5-sulphosalicylic acid in water). In polyethylene bottle, 100 ml
 

How to use Hungarian Red

On what surfaces can it be used

Hungarian Red is a dye (Acid Fuchsin) solution in water/acetic acid mixture that is used for staining fingerprints and footprints made in blood. Prints in blood are colored red after treatment with Hungarian red. Hungarian Red should not be used on absorbent surfaces like paper, carton materials, bed sheets, or carpet. It works very well on non-absorbent backgrounds like linoleum, glass, tiles, painted surfaces, or PVC floor covering.

Fix prints in blood before staining

Before staining, prints in blood should be fixed to prevent them from running (causing loss of detail) when the staining solution is applied. In general fixative is applied before any staining solution except Leuco Crystal Violet, which contains fixative.

How to fix the prints

Fixing is best done with a 2% solution of sulfosalicylic acid in water (20-23 gram in 1 liter). To make sure the print in blood is thoroughly fixed, we recommended the following method using absorbent paper (like filter paper, tissue paper, or paper towels) and a wash bottle containing sulfosalicylic acid solution.

Take a dry piece of the absorbent paper that is sized to cover the print(s). Hold it directly over the print-area, parallel to the surface and just slightly above the surface. To begin fixing, drop one edge of the paper to the surface and moisten heavily along its entire length, so that it is anchored and will stay in place. Starting from that edge, wet the paper progressively further while smoothing the wetted part onto the print and minimizing trapped air bubbles. Work carefully from one edge to the other. When possible, for example, with an object or with a print on an angled surface start from the top and work downwards.

Once the wet paper entirely covers the print, leave it there for a minimum of three minutes. When the blood is a thick layer, leave the paper there 5 minutes or more. Remove the paper. An excess of fixative can be rinsed away with water, but this is not necessary.

When the blood is relatively fresh, you will notice that fixing changes its color from dark red to dark brown.

Staining procedure

Once the print is fixed, it can be stained with Hungarian Red. The most economical way is using the sprayer. Spraying always causes some mist (aerosol) that will float around in the air. Especially when you stain large areas, do use some respiration protection. Alternatively, you can use a wash bottle (or with fingerprints even a disposable pipet).

After the staining solution is applied by sprayer or wash bottle, leave it in contact with the print for about one minute. Then wash the surface with water (or a 5% water/acetic acid mixture = 19:1 volume/volume ratio). If the print is located on a floor, the wash water might be removed with a vacuum cleaner that can handle water. Otherwise, remove it with paper towels or the like.

It is advantageous to immediately remove the water and water droplets from the processed area after rinsing, with compressed air or a powerful blower. If water droplets remain on the print, you will notice that some dye will dissolve in them. After drying that will be visible (possible loss of detail).

Lifting and fluorescence

Hungarian Red not only has a good staining capacity but has also some special characteristics. Stained prints can be lifted with a white BVDA gelatin lifter so that backgrounds are eliminated and/or parts on dark backgrounds become visible. The lifted print will also fluoresce, whereby details or prints that were hardly visible with the naked eye are now easily discernible in fluorescence.

Before lifting with a white gelatin lifter, the area should be completely dry. Be very careful not to trap air bubbles under the lifter surface and leave the lifter on for 15-30 minutes. Photograph within a couple of hours after lifting (the dye will slowly diffuse in and across the surface of the lifter, thereby blurring the print).

Lifting can be done more than once, with or without re-staining between lifts. Repeat lifting/staining in important cases.

Fluorescence is excited with green light (515-560 nm) and the filter used in front of the lens is a red filter (Kodak Wratten 25, a barrier filter of about 600 nm or a long-pass filter with cut-on of about 600 nm).

Safety

Hungarian Red is not labeled as a hazardous preparation. Since it is water based, no harmful fumes are liberated in use, although the acetic acid smell might be irritating. It will color the skin, although this will not last very long (in our experience less than a day). Contaminated clothing will be stain-free after regular washing.

Sulfosalicylic acid crystals are labeled as an irritant substance, the 2% solution is not labeled as a hazardous product. Given the fixative effect on blood, we strongly advise wearing gloves (vinyl gloves will suffice) when using it.

Avoid breathing aerosols (mists) of both the fixative and Hungarian Red. Likewise avoid contact with solutions and wash your hands after each session, especially before eating or drinking.

Effect on DNA analysis

According to a Canadian study useable DNA can still be retrieved from treated prints after staining with Hungarian Red.


Name: Acid Fuchsin, Acid Violet 19
CAS-No.: 3244-88-0
 

Coomassie Blue

Main uses: Blood traces on non-porous surfaces
 
Coomassie Blue, a reagent like Amido Black that stains proteins, is methanol-based. Traces in blood are stained blue to purple.
 
Product #
Description
Quantity
Add
 
 
B-88700
Coomassie Blue, 500 ml in a spray bottle
 


Name: Coomassie Brilliant Blue R, CI Acid Blue 83, Benzenemethanaminium, N-(4-((4-((4-ethoxyphenyl)amino)phenyl)(4-(ethyl((3-sulfophenyl)methyl)amino)phenyl)methylene)-2,5-cyclohexadien-1-ylidene)-N-ethyl-3-sulfo-, inner salt, sodium salt (1:1)
CAS No.:
6104-59-2
 

Leuco Crystal Violet

LCV (Leuco Crystal Violet), also known as ALCV (Aqueous Leuco Crystal Violet), is a coloring reagent for blood that is based on the blood-catalyzed reaction of hydrogen peroxide with LCV, whereby the colorless LCV is oxidized to the purple crystal violet (the same dye as used in Gentian Violet). Because this oxidation will also occur slowly under the influence of light and oxygen, the contrast of the visualized traces and the background is not permanent. After a period of time the background will also be colored purple.
To maximize the shelf-life, the hydrogen peroxide is separately packaged in an opaque, brown plastic bottle. Before use, the hydrogen peroxide is added to the other solution (in a mixing ratio of 1:4). Both solutions are water-based. The LCV solution already contains fixative, so there is no need to fix blood before staining.
In contrast to other staining solutions, LCV is very suitable for porous surfaces.
 

Product #
Description
Quantity
Add
 
 
B-88600
Leuco Crystal Violet set 500 ml
 

Leuco Crystal Violet (LCV) for staining of marks in blood

Leuco Crystal Violet (LCV) is a reagent for the detection and staining of marks in blood. It is a colorless to light-blue water-based (400 ml in a brown glass bottle, solution A) that is mixed just before use with the included 15% hydrogen peroxide solution (100 ml in a brown plastic bottle). The mixing ratio is 4 parts solution A to 1 part peroxide solution (B).


The reagent is sprayed over the surface where marks in blood are suspected. When the (mixed) reagent comes in contact with blood the hydrogen peroxide is broken down by the hemoglobine. In turn the oxidized hemoglobin oxidizes the the colorless LCV to strongly purple-colored crystal violet. The hemoglobin is now back in it's original state and can be oxidized by hydrogen peroxide again. This is therefore a catalytic reaction (the hemoglobin is the catalyst).


Leuco Crystal Violet reacts to Crystal Violet

Crystal violet is also known as gentian violet and Basic Violet 3. On non-porous surfaces rinsing with demineralized water to remove excess reagent (which will slowly become purple due to oxidation by air helped by light of the unreacted LCV) is an option.


Once the oxidation to crystal violet has occurred, removing the purple stain is often cumbersome if not impossible. This should be kept in mind when using it on crime scenes.

Advantages of the reagent are:


  • in principle there is no need for rinsing which makes it less labor intensive than for instance dye solutions like Amido Black and Hungarian red and makes it possible to use it on materials that do not lend themselves to rinsing like carpet.
  • it can be used both on porous and non-porous surfaces.
  • the marks do not need prior fixing because the reagent already contains fixative (the working solution contains 2% sulfosalicylic acid).
  • it does not contain volatile solvents and with respect to chemical composition not dangerous.

After the application of LCV protein stains like Amido Black and Acid Violet 17 can still be used. Apparently, LCV can also be used after luminol [1], however it first needs fixing with 2% sulfosalicylic acid otherwise leaching with loss of detail occurs.


Under the influence of light unreacted LCV can still be oxidised causing background coloration. This process is however very slow so there is plenty of time to photographically record visualized marks.

Crystal violet is known [2] to fluoresce in the deep-red and infrared after oxidation (especially at low concentrations.) This might yield additional marks or make it possible to enhance developled marks.

Shelf life

Once solution A and B are mixed the shelf life when stored in the dark and cold (refrigerator) is several months. To maximize shelf life BVDA ships the reagent as two separate solutions, one of which contains the hydrogen peroxide. This extends the shelf life appreciably although an exact time is hard to name. It might also depend on the particular batch of LCV and/or other small variations in the chemicals used. On standing the LCV solution slowly darkens. When the surface already becomes purple when there is no blood present, the reagent should obviously be discarted. The hydrogen peroxide solution also slowly decomposes.


Therefore, we strongly recommend to store the reagent in the refrigerator.

History

LCV was conceived and over the course of several years improved by John F. Fischer, who worked for the Orange County Sheriff's Office in Orlando (Florida). The FBI lab in Washington started using it at the end of 1993.
In June 1994 John Fisher gave a presentation on LCV at the FBI academy in Quantico (International Symposium on Footwear and Tire Tread Impression Evidence). Bill Bodziak who at the time still worked for the FBI gave a lecture on it at the first SPTM-conference (the working group later became part of ENFSI) in Helsinki (Finland), May 1995 [1].
In 1996 Bill Bodziak published an article on LCV in the journal Forensic Science International [3].

Footnotes

[1]  Handout of a lecture by Bill Bodziak: "The use of Leuco Crystal Violet to enhance shoe prints in blood", Shoe Print/Tool Mark Examiners (SPTM) conference, May 1995, Helsinki (Finland).
[2] Bramble, K.; Cantu, A.A.; Ramotowski, R.R.; Brennan, J.S., "Deep Red to Near Infrared (NIR) Fluorescence of Gentian Violet-treated Latent Prints", Journal of Forensic Identification 2000, Vol. 50, no. 1, pp. 33-49.
Also see the letter by Menzel, E.  Journal of Forensic Identification 2000, Vol. 50, No. 3, pp. 245-249.
[3]  Bodziak, William J. "Use of leuco crystal violet to enhance shoeprints in blood", Forens. Sci. Int., 1996, Vol. 82, pp. 45-52, [doi:10.1016/0379-0738(96)01965-2]