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Lab Recipes; Buffer and Reagents

Lab Recipes; Buffers and Reagents

A

Acrylamide (30%)

Acrylamide (40%)

Alkaline Lysis Buffer A

Alkaline Lysis Buffer B

Alkaline Lysis Buffer C

B

B5 Fixative

Bouin's Fixative

Bradford's Reagent

Bromthymol Blue

C

Cell Staining Buffer

Coomassie Blue


D

Denhardt solution (50x)

Denhardt solution (100x)

DEPC treated water

Destain Solution

DNA Loading Buffer (Orange G)

DNA Loading Buffer (XC/BB)

E

EDTA

Ehrlich's Solution

ELISA Blocking Solution

ELISA Coating Buffer

F

Field's Stain

Flagella Stain

Formaldehyde Gel Running Buffer


G

Gentian Violet Stain

Giemsa Stain

Glycerol Tolerant Gel Buffer (GTGB) 20x

H

Hellings (10x)

Hollande's Fixative

K

Kovacs' Reagent


L

Laemmli Sample Buffer (2x)

M

Modified RIPA Buffer

N

NADI Reagent

NP-40 Cell Lysis Buffer


P

Papanicolaou's Stain

Paraformaldehyde (2%)

Pepsin

Phosphate Buffered Saline (PBS) 1x

Phosphate Buffered Saline (PBS) 10x

Phosphate Buffered Saline Tween-20 (PBST) 1x

Proteinase K Solution

R

Red Blood Cell (RBC) Lysis Buffer

S

Schiff's Reagent

Sodium Citrate Buffer

SSPE 20X

STET

Synthetic Sea Water


T

TAE 50x

TBE 10x

TE 1x

TES

Tissue Fixation Solution

Tissue Homogenization Buffer for ELISA

TNE 1x

TPE 1x

Tris Glycine Buffer 5x

Tris-Buffered Saline (TBS)

Tris-Buffered Saline Tween-20 (TBST)

Trypan Blue

TTE 1x

W

Western Blotting Transfer Buffer

Wolfe's Mineral Solution

Wolfe's Vitamin Solution

X

X-gal staining solution


Z

Zenker's Fixative


Acrylamide (30%)

  1. Dissolve 290g of acrylamide and 10g of N,N'-methylbisacrylamide in 600ml of H2O. Heating may be necessary to dissolve the acrylamide.
  2. Adjust the volume to 1L with H2O.
  3. Sterilize the solution by filtration (0.45 micron pore size).
  4. Check the pH (should be 7.0 or less).
  5. Store in dark bottles at room temperature.

Caution: Acrylamide is a neurotoxin. Use extreme caution when handling anything with acrylamide and bisacrylamide. Protective equipment should be worn.


Acrylamide (40%)

  1. Dissolve 380g of acrylamide and 20g of N,N'-methylbisacrylamide in 600ml of H2O. Heating may be necessary to dissolve the acrylamide.
  2. Adjust the volume to 1L with H2O.
  3. Sterilize the solution by filtration (0.45 micron pore size).
  4. Check the pH (should be 7.0 or less).
  5. Store in dark bottles at room temperature.

Caution: Acrylamide is a neurotoxin. Use extreme caution when handling anything with acrylamide and bisacrylamide. Protective equipment should be worn.


Alkaline Lysis Buffer A

  1. Add the following to 100ml distilled H2O
    • 5ml of 20% SDS
    • 2ml of 10M NaOH

Alkaline Lysis Buffer B

  1. Add the following to 100ml distilled H2O
    • 29.4g KAc
    • 11.5ml of Glacial Acetic Acid

Alkaline Lysis Buffer C

  1. Add the following to 900ml distilled H2O
    • 9g Glucose
    • 3g Tris
    • 20ml of 0.25M EDTA
  2. q.s. to 1000ml with distilled H2O
  3. Autoclave using standard conditions
  4. Allow to cool to room temperature
  5. Store at 4oC

When ready to use add 0.02g lysozyme to 100ml of alkaline lysis buffer C to yield alkaline lysis buffer A.


B5 Fixative

  1. Stock solution (stable for approximately 1 year). Mix well and do not use any metal utensils or foil lined lids.
    • 12.0g Mercuric chloride
    • 2.5g Sodium acetate
    • 200ml Distilled water 200 ml
  2. Working solution (prepare right before use)
    • 20ml of stock solution
    • 2ml of formaldehyde

Caution: Mercuric chloride is a skin and eye irritant and Formaldehyde is a carcinogen. Use protective equipment and prepare under hood in a well ventilated area. Reference proper procedures and disposal.


Bouin's Fixative

  1. Stock solution
    • 3L Saturated picric acid
    • 1L Formaldehyde
    • 200ml Glacial acetic acid

Caution: Picric acid can become explosive if allowed to dry out and is toxic through skin exposure. Formaldehyde is a carcinogen. Glacial acetic acid is caustic. Use protective equipement and prepare under hood in a well ventilated area. Reference proper procedures and disposal.


Bradford's Reagent

  1. Dissolve 50mg of Coomassie Blue G250 in 50ml of methanol
  2. Add 100ml of 85% H3PO4 to the solution from step 1
  3. Add the solution from step 2 into 500ml of H2O and mix
  4. Filter to remove and precipitates
  5. Add an additional 350ml of H2O
  6. Store at 4oC.

Caution: Always add acid slowly into water and do not add water into acid.


Bromthymol Blue

Aqueous version

  1. Add 0.1g Bromthymol blue into 16ml of 0.01N NaOH
  2. Mix in a mortar or small tube
  3. Dilute to 250ml with distilled water
  4. Use 5 drops in 10ml of test solution

Alcohol version

  1. Add 0.5g of Bromthymol blue into 500ml of 95% ethanol and dissolve
  2. Add 500ml of distilled water
  3. Filter and store at room temperature

 


Cell Staining Buffer

  1. Dissolve 8g of NaCl, 0.2g of KCl, 1.44g of Na2HPO4, and 0.24g of KH2PO4 in 800ml distilled H2O.
  2. Add 20 ml of heat inactivated FBS.
  3. Add 0.9 grams of sodium azide.
  4. Adjust pH to 7.4 with HCl.
  5. Adjust volume to 1L with additional distilled H2O.
  6. Sterilize by filtration.
  7. Store at 4oC.

Coomassie Blue Solution

  1. Dissolve 2g Coomassie Blue (Serva Blau) in 250ml water
  2. Slowly add 75ml of glacial acetic acid
  3. Add 500ml of ethanol
  4. q.s. to 1000ml with water

Final concentrations:
0.2% Coomassie Blue
7.5% Acetic Acid
50% Ethanol


Denhardt solution (50x)

  1. Add the following to 900ml distilled H2O
    • 10g Ficoll 400
    • 10g Polyvinilpyrolidone
    • 10g BSA
  2. Filter solution prior to storage through a 0.2uM filter
  3. Store at 4oC
  4. Warm to appropriate temperature prior to use

Denhardt solution (100x)

  1. Add the following to 900ml distilled H2O
    • 20g Ficoll 400
    • 20g Polyvinilpyrolidone
    • 20g BSA
  2. Filter solution prior to storage through a 0.2uM filter
  3. Store at 4oC
  4. Warm to appropriate temperature prior to use

DEPC Treated Water

  1. Add 1ml of 0.1% Diethylpyrocarbonate (DEPC) to 1000ml distilled water
  2. Mix well and let set at room temperature for 1 hour
  3. Autoclave
  4. Let cool to room temperature prior to use

Destain Solution

  1. Add the following to 800ml distilled H2O
    • 7.5ml of Glacial Acetic Acid
    • 100ml of Ethanol
  2. q.s. to 1000ml with distilled H2O

DNA Loading Buffer (Orange G)

  1. Dissolve 20g of Sucrose in 40ml water
  2. Dissolve 100mg of Orange G in above solution
  3. q.s. to 50ml with water

EDTA

(1mM EDTA, 0.05% Tween 20, pH 8.0):
EDTA 0.37 g
Distilled water 1000 ml
Mix to dissolve.
Adjust pH to 8.0 using 1N NaOH.
Then add 0.5 ml of Tween 20 and mix well.

Ehrlich's Solution

  1. Dissolve 1g of Para-dimethylamino benzaldehyde in 95ml of 95% ethanol
  2. Add 20 ml of concentrated HCl to the solution from step 1
  3. Mix well

Caution: Always add acid slowly into water and do not add water into acid.


ELISA Blocking Solution

  1. Dissolve 5.3g of Na2CO3 in 900ml distilled H2O
  2. Dissolve 4.2g of NaHCO3 in the solution from step 1
  3. Dissolve 1g sodium azide in the solution from step 2
  4. Dissolve 5g BSA in solution from step 3
  5. Alternatively you can also use 50g non-fat dry milk for blocking solution
  6. pH to 9.6
  7. Adjust volume to 1L with additional distilled H2O

ELISA Coating Buffer

  1. Dissolve 5.3g of Na2CO3 in 900ml distilled H2O.
  2. Dissolve 4.2g of NaHCO3 in the solution from step 1.
  3. Dissolve 1g sodium azide in the solution from step 2.
  4. pH to 9.6.
  5. Adjust volume to 1L with additional distilled H2O.

Field's Stain

Field's Solution #1

  1. Dissolve 1.6 g of methylene blue in 1 liter of distilled water
  2. Dissolve 2.6g of Na2HPO4 (anhydrous) to the solution from step 1
  3. Dissolve 1g of Azure 1 in the solution from step 2
  4. Dissolve 2.6 g of KH2PO4 in the solution from step 3
  5. Place on mild heat with stirring or shaking for 45 minutes to 1 hour
  6. Let stand at room temperature for 24 hours
  7. Filter

Field's Solution #2

  1. Dissolve 2g of Eosin Y in 1 liter of distilled water
  2. Dissolve 2.6g of Na2HPO4 in the solution from step 1
  3. Dissolve 2.6g of KH2PO4 in the solution from step 2
  4. Filter

Flagella Stain

  1. Dissolve 20g of tannic acid in 30ml of 60% ethanol
  2. Dissolve 36g of aluminum chloride (hydrated) to the solution from step 1
  3. Dissolve 20g of ZnCl2 in the solution from step 2
  4. Dissolve 3g of rosaniline HCl in the solution from step 3
  5. q.s. to 70ml with 60% ethanol

Formaldehyde Gel Running Buffer

  1. Add the following to 800ml distilled H2O
    • 209.27 g MOPS
    • 68.04 g AcONa x 3H2O
    • EDTA x 2H2O
  2. Adjust pH 7.0 using 10N NaOH
  3. q.s. to 1 liter with distilled H2O
  4. Filter sterilize through 0.2 µm filter

Gentian Violet Stain

  1. Dissolve 1g gentian violet in 20ml of 95% ethanol
  2. Add 60ml 5% formalin to the solution from step 1 and mix completely
  3. Filter solution to remove any precipitate

 


Giemsa Stain

Stock Solution

  1. Dissolve 3.8g of Giemsa powder into 250ml of methanol
  2. Heat the solution from step 1 to ~60oC
  3. Slowly add in 250ml of glycerin to the solution from step 2
  4. Filter the solution from step 3
  5. The solution needs to stand a period of time prior to use. Although times vary based on who you ask a minimum of two months is usually recomended

Working Solution

  1. Add 10ml of stock solution to 80ml of distilled water and 10ml of methanol

Glycerol Tolerant Gel Buffer (GTGB) 20X

  1. Dissolve in 750 ml of H2O:
    • 216g Tris base
    • 72g Taurine (C2H7NO3S)
    • 4g of Na2EDTAx2H2O
  2. After solid is dissolved, adjust volume to 1L with H2O

Hellings (10X)

  1. Dissolve the following in 800ml distilled H2O
    • 60.55g Tris
    • 16.5g NaAc (anhydrous)
    • 6.95g EDTA
  2. Adjust pH to 8.05 with glacial acetic acid
  3. qs to 1000ml with distilled H2O

Hollande's Fixative

Stock solution

  • 75g Copper acetate (cupric)
  • 3L Distilled water
  • 120g Picric acid
  • 300ml Formaldehyde
  • 45ml Glacial acetic acid

Caution: Picric acid can become explosive if allowed to dry out and is toxic through skin exposure. Formaldehyde is a carcinogen. Glacial acetic acid is caustic. Use protective equipement and prepare under hood in a well ventilated area. Reference proper procedures and disposal.


Kovacs' Reagent

  1. Add 25ml of concentrated HCl to 75ml of Amyl alcohol
  2. Dissolve 5g of paradimethylamino-benzaldehyde in the solution from step 1
  3. Aliquot to single use vials
  4. Store at 4oC in closed vials

Laemmli Sample Buffer (2X)

4% SDS
20% glycerol
10% 2-mercaptoethanol (or DTT) (add immediately before use)
0.004% bromphenol blue
0.125M Tris-Cl, pH 6.8

Modified RIPA Buffer

10 mM Tris, pH 7.4
100 mM NaCl
1 mM EDTA
1 mM EGTA
1 mM NaF
20 mM Na4P2O7
2 mM Na3VO4
1% Triton X-100
10% glycerol
0.1% SDS
0.5% deoxycholate
plus protease inhibitors: Aprotinin, leupeptin, pepstatin: 1ug/ml each
Add 1mM PMSF immediately before use.


See:
C2605-70 - Cell Extraction Buffer
P9070 - Protease Inhibitor Cocktail


NP-40 Cell Lysis Buffer

50mM Tris-HCl pH 8.0
150mM NaCl
1% NP-40
plus protease inhibitors: Aprotinin, leupeptin, pepstatin: 1ug/ml each
Add 1mM PMSF immediately before use.

See:
P9070 - Protease Inhibitor Cocktail

 


NADI Reagent

Solution # 1

  • 1% alpha-naphthol in 95% ethanol

Solution # 2

  • 1% N,N-dimethyl-p-phenylenediamine HCl in water

Solutions 1 and 2 are mixed in equal volumes just prior to use.


Papanicolaou's Stain

  1. Mix 11.25ml of a 10% eosin Y solution into 450ml of 100% ethanol.
  2. Mix 2.25ml of a 10% aqueous solution of light green SF into the solution from step 1.
  3. Mix 25ml of a 1% bismarck brown solution (in ethanol) into the solution from step 2.
  4. Mix 10ml of a 10% aqueous solution of phosphotongstic acid into the solution from step 3.
  5. Mix 0.3ml of an aqueous solution of saturated lithium carbonate into the solution from step 4.

Paraformaldehyde (2%)

To make 100ml of 2% Paraformaldehyde

  1. Add 2 grams of paraformaldehyde to 48 ml of water
  2. Heat to dissolve
  3. Add NaOH dropwise until solution clears (10-20 drops of 2M)
  4. Add 50ml of 2x PBS and mix
  5. Remove from heat and place on ice
  6. pH from 7.2 to 7.4

Pepsin

Pepsin Stock Solution (1% in 10mM HCl):

Pepsin 100 mg
10mM HCl (pH 2.0) 10 ml
Mix to dissolve. Store at -20 ºC


Pepsin Working Solution (0.5% in 5mM HCl):

Pepsin Stock Solution (1%) 1 ml
Distilled water 1 ml
Mix well.


Phosphate Buffered Saline (PBS) 1x

  1. Dissolve the following in 800ml distilled H2O.
    • 8g of NaCl
    • 0.2g of KCl
    • 1.44g of Na2HPO4
    • 0.24g of KH2PO4
  2. Adjust pH to 7.4.
  3. Adjust volume to 1L with additional distilled H2O.
  4. Sterilize by autoclaving

Phosphate Buffered Saline (PBS) 10x

  1. Dissolve the following in 800ml distilled H2O.
    • 80g of NaCl
    • 2.0g of KCl
    • 14.4g of Na2HPO4
    • 2.4g of KH2PO4
  2. Adjust pH to 7.4.
  3. Adjust volume to 1L with additional distilled H2O.
  4. Sterilize by autoclaving

Phosphate Buffered Saline Tween-20 (PBST) 1x

  1. Dissolve the following in 800 ml of distilled H2O
    • 8g of NaCl
    • 0.2g of KCl
    • 1.44g of Na2HPO4
    • 0.24g of KH2PO4
    • 2ml of tween-20
  2. Adjust pH to 7.2
  3. Adjust volume to 1L with additional distilled H2O
  4. Sterilize by autoclaving

Proteinase K Solution

(20 ug/ml in TE Buffer, pH 8.0):
TE Buffer (50mM Tris Base, 1mM EDTA, 0.5% Triton X-100, pH 8.0):

Tris Base 6.10 g
EDTA 0.37 g
Triton X-100 5 ml
Distilled water 1000 ml

Mix to dissolve. Adjust pH 8.0 using concentrated HCl (10N HCl). Store at room temperature.


Red Blood Cell (RBC) Lysis Buffer

  1. Dissolve the following in 800ml distilled H2O
    • 8.3g NH4Cl
    • 1.0g KHCO3
    • 1.8ml of 5% EDTA
  2. Filter sterilize through 0.2um filter
  3. qs to 1000ml with distilled H2O

Red Blood Cell (RBC) Lysis Buffer

  1. Dissolve the following in 800ml distilled H2O
    • 8.3g NH4Cl
    • 1.0g KHCO3
    • 1.8ml of 5% EDTA
  2. Filter sterilize through 0.2um filter
  3. qs to 1000ml with distilled H2O

Schiff's Reagent

  1. Dissolve 5g of basic fuchsin in 900ml of boiling distilled water
  2. Cool to approximately 50oC and slowly add 100ml of 1N HCl
  3. Cool to approximately 25oC and dissolve 10g of K2S2O5
  4. Shake for 3 minutes and incubate in the dark at room temperature for 24 hours
  5. Add 5 grams of fine activated charcoal and shake for 3 minutes
  6. Filter solution (should be clear)
  7. Store at 4oC in a foil covered bottle

Sodium Citrate Buffer

(10mM Sodium Citrate, 0.05% Tween 20, pH 6.0):

Tri-sodium citrate (dihydrate) 2.94 g
Distilled water 1000 ml

Mix to dissolve. Adjust pH to 6.0 with 1N HCl and then add 0.5 ml of Tween 20 and mix well


SSPE 20X

  1. Dissolve the following in 800ml of distilled H2O.
    • 175.3g of NaCl
    • 27.6g of NaH2PO4-H2O
    • 7.4 g of EDTA
  2. Adjust the pH to 7.4 with 10N NaOH.
  3. Adjust the volume to 1L with additional distilled H2O.
  4. Sterilize by autoclaving.

STET

  1. Dissolve the following in 20ml water
    • 4.0 g of sucrose
    • 2.5ml of Triton X-100
    • 200ul of 0.25M EDTA
    • 2.5ml of 1M Tris-HCl
    • 50ul of 20% Sodium Azide
  2. q.s. to 50ml with water

Synthetic Sea Water

  1. Dissolve the following in 850ml of distilled water
    • 24g NaCl
    • 11g MgCl2-6H2O
    • 4g Na2SO4
    • 2g CaCl2-6H2O
    • 0.7g KCl
    • 0.1g KBr
    • 0.03g H3BO3
    • 0.005g NaSiO3-9H2O
    • 0.04g SrCl2-6H2O
    • 0.003g NaF
    • 0.002g NH4NO3
    • 0.001g Fe3PO4-4H2O
  2. Adjust pH to 7.8
  3. Bring to 1000ml with distilled water
  4. Autoclave or filter sterilize

TAE 50x

  1. Add the following to 900ml distilled H2O
    • 242g Tris base
    • 57.1ml Glacial Acetic Acid
    • 18.6 g EDTA
  2. Adjust volume to 1L with additional distilled H2O

TBE 10x

  1. Add the following to 800ml H2O
    • 108g Tris base.
    • 55g Boric acid
    • 9.3g EDTA
  2. Adjust volume to 1L with additional distilled dH2O

TE 1x

  1. Add the following to 990ml distilled H2O
    • 10ml  1M Tris-HCl (pH 8.0)
    • 400µl  0.25 M EDTA

TES

  1. Add 5ml of 1M Tris pH 8.0 to 50ml distilled water
  2. Add 2ml of 0.25M EDTA
  3. Dissolve 0.3g NaCl
  4. q.s. to 100ml with distilled water

Tissue Fixation Solution

To 400ml of PBS add:

  1. 100ml of 10% Formalin
  2. 200ul of gluteraldehyde
  3. 100ul NP-40

Tissue Homogenization Buffer for ELISA

Stock Solution:

  1. Weigh out 0.025 grams of Aprotinin and Luepeptin and dissolve in 25ml of water
  2. Mix and aliquot into 70ul single use aliquots

To prepare homogenization buffer

  1. Add 50ul of stock solution to 100ml of PBS
  2. Sterilze by filtration

Final concentration is 0.5ug/ml

 


TNE 1x

  1. Add the following to 800ml of distilled H2O
    • 12.1g Tris base
    • 3.7g Na2EDTA.2H2O
  2. Adjust pH to 7.4 with concentrated HCl
  3. Adjust volume to 1L with additional distilled H2O

TPE 1x

  1. Add the following to 900ml distilled H2O
    • 108 Tris base
    • 15.5ml 85% Phosphoric acid
    • 40ml of 0.5M EDTA (pH 8.0)
  2. qs to 1000ml with distilled H2O

Tris Glycine Buffer 5x

  1. Dissolve in 700 ml of H2O:
    • 15.1g Tris base
    • 94g glycine
    • 50ml of 10% SDS
  2. After solid is dissolved, adjust volume to 1L with H2O

Tris-Buffered Saline (TBS)

  1. Dissolve the following in 800ml of distilled H2O.
    • 8g of NaCl, 0.2g of KCl
    • 3g of Tris base
  2. Add 0.015g of phenol red.
  3. Adjust the pH to 7.4 with HCl.
  4. Add distilled H2O to 1L.
  5. Sterilize by autoclaving.

Tris-Buffered Saline Tween-20 (TBST)

  1. Dissolve the following in 800ml of distilled H2O
    • 8.8 g of NaCl
    • 0.2g of KCl
    • 3g of Tris base
  2. Add 500ul of Tween-20
  3. Adjust the pH to 7.4
  4. Add distilled H2O to 1L
  5. Sterilize by filtration or autoclaving

Trypan Blue

  1. Dissolve the following in 80ml PBS
    • 0.4g Trypan blue
  2. Bring to a slow boil
  3. Cool to room temperature
  4. qs to 100ml with PBS

Add 0.1 volumes of trypan blue solution to 0.9 volumes of cells in PBS. Unstained cells are viable.


TTE 1x

  1. Add the following to 988ml distilled H2O
    • 10ml  1M Tris-HCl (pH 8.0)
    • 400µl  0.25 M EDTA
    • 2ml Triton X-100

Western Blotting Transfer Buffer

  1. Add the following to 800ml H2O
    • 36.35g Tris
    • 150g Glycine
    • 4g SDS
  2. q.s. to 1000ml with distilled H2O

Wolfe's Mineral Solution

  1. Add 1.5g nitrilotriacetic acid to approximately 500 ml of water and adjust to pH 6.5 with KOH to dissolve.
  2. Add the following:
    • 3.0g MgSO4-7H2O
    • 0.5g MnSO4-H2
    • 1.0g NaCl
    • 0.1g FeSO4-7H2O
    • 0.1g CoCl2-6H2O
    • 0.1g CaCl2
    • 0.1g ZnSO4-7H2O
    • 0.01g CuSO4-5H2O
    • 0.01g AlK(SO)4-12H2O
    • 0.01g H3BO3
    • 0.01g Na2MoO4-2H2O
  3. q.s. to 1000ml
  4. Filter sterilize

Wolfe's Vitamin Solution

  1. Dissolve (or add approriate concentrations solutions) for the following into 900ml of distilled water
    • 10mg Pyridoxine hydrochloride
    • 5.0mg Thiamine-HCl
    • 5.0mg Riboflavin
    • 5.0mg Nicotinic acid
    • 5.0mg Calcium D-(+)-pantothenate
    • 5.0mg p-Aminobenzoic acid
    • 5.0mg Thioctic acid
    • 2.0mg Biotin
    • 2.0mg Folic Acid
    • 0.1mg Vitamin B12
  2. q.s. to 1000ml
  3. Filter sterilize

X-gal staining solution

  1. Add the following to 495ml of PBS
    • 1.050g Ferrocyanide
    • 0.825g Ferricyanide
    • 0.050g NaDeoxycholate
    • 0.2g MgCl2
    • 500ul Nonidet-P40
    • 0.5g X-gal in 5ml N,N-Dimethylformamide

Xilene Cyanol/Bromophenol Blue DNA Loading Buffer 10x

  1. Dissolve in 6.25 ml of H2O
    • .025g of Xilene cyanol
    • .025g of Bromophenol Blue
    • 1.25ml of 10% SDS
    • 12.5ml of glycerol

Zenker's Fixative

  1. Stock solution
    • 50g Mercuric chloride
    • 25g Potassium dichromate
    • 10g Sodium sulfate
    • 1L Distilled water
  2. Working solution
    • 95ml Zenker’s stock
    • 5ml Glacial acetic acid

Caution: Mercuric chloride is a skin and eye irritant. Potassium dichromate is toxic by inhalation of dust and ingestion. Glacial acetic acid is caustic. Use protective equipement and prepare under hood in a well ventilated area. Reference proper procedures and disposal.