Products

Basement Membrane

Basement Membrane Stained Histology Slide

Validation Stain:  Jones Basement Membrane
Other Applicable Stains:  Gomori, PAMM

 

Tissue: 
(+) Kidney
Adhesive or Charged Slides: 
Superfrost +

Product Options:

Part # 4055A 10/set $60.80
Part # 4055B 98/set $452.40

PRODUCT SPECIFICATIONS:

Tissue:  Positive staining kidney.
Fixation: Formalin 10%, Phosphate Buffered (Part 1090).
Section/Glass: Paraffin sections cut at 4 microns on Superfrost™ Plus slides.
Quality Control Stain:  Jones Basement Membrane quality control stained slide(s) included.
Reactivity: Guaranteed product specific reactivity for one year from date of receipt. Revalidate after one year to verify continued reactivity. 
Storage: 15-30°C in a light deprived and humidity controlled environment.
Intended Use: To verify histological techniques and reagent reactivity.

Before using unstained control slides, review the enclosed stained slide(s) to ensure that this tissue source is acceptable for testing needs.

 

CONTROL SLIDE VALIDATION:                                                         

With Jones Basement Membrane Stain Kit Part 9167A Individual Stain Solution
Solution A: Methenamine 3%, Aqueous 250 ml Part 12239
Solution B: Silver Nitrate 5%, Aqueous 50 ml Part 13805
Solution C: Sodium Borate 5%, Aqueous 50 ml Part 13826
Solution D: Periodic Acid 0.5%, Aqueous 250 ml Part 13308
Solution E: Gold Chloride 0.25%, Aqueous 250 ml Part 11287
Solution F: Sodium Thiosulfate 2.5%, Aqueous 250 ml Part 13889
Solution G: Light Green SF Yellowish Stain 0.1%, Aqueous 250 ml Part 12203

 

APPLICATION:

Newcomer Supply Basement Membrane Control Slides are for positive histochemical staining of basement membranes in tissue sections.

 

PRESTAINING PREPARATION:

    1. Heat dry sections in oven according to your laboratory protocol.
    2. All glassware/plasticware must be acid cleaned prior to use.
        1. See Procedure Notes #1 and #2.
    3. Prepare Silver-Methenamine Working Solution and mix well:
        1. Solution A: Methenamine 3%, Aqueous      40 ml
        2. Solution B: Silver Nitrate 5%, Aqueous 2 ml
        3. Solution C: Sodium Borate 5%, Aqueous 4 ml
    4. Preheat Silver-Methenamine Working Solution to 45°-60°C in a water bath 20-30 minutes before use.
        1. Maintain solution between 45°-60°C to minimize precipitate.
        2. Do not preheat solution if using Microwave Modification.

 

NEWCOMER SUPPLY VALIDATION PROCEDURE:

    1. Deparaffinize sections thoroughly in three changes of xylene, 3 minutes each. Hydrate through two changes each of 100% and 95% ethyl alcohols, 10 dips each.  Wash well with distilled water.
        1. See Procedure Notes #3 and #4.
    2. Place in Solution D: Periodic Acid 0.5%, Aqueous for 15 minutes.
    3. Wash in tap water for 5 minutes; rinse in distilled water.
    4. Incubate slides in preheated Silver-Methenamine Working Solution (Step #4) at 45°-60°C oven or at room temperature for 12-18 minutes until sections appear paper-bag brown.
    5. Periodically remove control; rinse in warm distilled water, check microscopically for adequate silver impregnation. Basement membranes should be dark brown. If tissue structures are not sufficiently dark, return slides to warm silver solution.  Recheck at 2-3 minute intervals until desired intensity is achieved.
        1. Staining at room temperature will require longer incubation.
    6. Microwave Modification: See Procedure Note #5.
        1. Place sides in a plastic Coplin jar with prepared Silver-Methenamine Working Solution (Step #3). Microwave for 3 minutes at 70°C.
        2. Check microscopically for adequate development.
        3. If additional incubation is required, return slides to heated silver solution and recheck at regular intervals.
    1. Rinse in three changes of distilled water.
    2. Tone in Solution E: Gold Chloride 0.25%, Aqueous for 1 minute.
    3. Rinse well in three changes of distilled water.
    4. Place in Solution F: Sodium Thiosulfate 2.5%, Aqueous; 2 minutes.
    5. Wash in tap water for 5 minutes; rinse in distilled water.
    6. Counterstain in Solution G: Light Green SF Yellowish Stain 0.1%, Aqueous for 1 minute.
    7. Quickly rinse slides in two changes of distilled water.
    8. Dehydrate in two changes each of 95% and 100% ethyl alcohol. Clear in three changes of xylene, 10 dips each; coverslip with compatible mounting medium.

 

RESULTS:

Kidney glomerular basement membranes Black
Intra-glomerular deposits Black
Reticular Fibers Black
Nuclei Outlined in black
Background Light Green

 

PROCEDURE NOTES:

    1. Acid clean all glassware/plasticware (Part 12086) and rinse thoroughly in several changes of distilled water.
    2. No metals of any kind should come in contact with silver solutions to prevent precipitation of silver salts.  Use plastic forceps (Part 5500) or paraffin coated metal forceps.
    3. Drain slides after each step to prevent solution carry over.
    4. Do not allow sections to dry out at any point during procedure.
    5. The microwave procedure was tested using a laboratory-grade microwave oven. This procedure is a guideline and techniques should be developed for use in your laboratory.
    6. If using a xylene substitute, follow manufacturer’s recommendation for deparaffinization and clearing steps.

 

REFERENCES:

    1. Jones, David B. “Nephrotic Glomerulonephritis,” American Journal of Pathology2 (1957): 313–329.
    2. Luna, Lee G. Manual of Histologic Staining Methods of the Armed Forces Institute of Pathology. 3rd ed. New York: Blakiston Division, McGraw-Hill, 1968. 97-99.
    3. Sheehan, Dezna C. and Barbara B. Hrapchak. Theory and Practice of Histotechnology. 2nd ed. St. Louis: Mosby, 1980. 187-188.
    4. Modifications developed by Newcomer Supply Laboratory.