Gram Positive & Gram Negative Bacteria, Animal

Gram (+) and Gram (-) In animal tissue with specific infection or disease.

Tissue: 
(+) Animal Tissue / (-) Lung
Adhesive or Charged Slides: 
Superfrost +

Product Options:

Part # 4254A 10/Set $52.00
Part # 4254B 98/Set $365.00

 

PRODUCT SPECIFICATIONS:

Tissue:  Positive staining animal tissue and negative staining human lung.

Fixation: Formalin 10%, Phosphate Buffered (Part 1090).

Section/Glass: Paraffin sections cut at 4 microns on Superfrost™ Plus slides.

Quality Control Stain:  Brown-Hopps 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 Gram, Brown-Hopps Stain Kit: Part 9124A Individual Stain Solution
Solution A:  Crystal Violet Stain 1%, Aqueous, Brown-Hopps 250 ml Part 1041
Solution B:  Iodine, Gram, Aqueous 250 ml Part 1140
Solution C:  Basic Fuchsin Stain 0.25%, Aqueous 250 ml Part 1011
Solution D:  Gallego Solution 250 ml Part 1098
Solution E:  Picric Acid-Acetone 0.05% 250 ml Part 13351
Solution F:  Acetone-Xylene 1:1 250 ml Part 10015
Acetone, ACS Part 10014

 

APPLICATION:

Newcomer Supply Gram Positive & Gram Negative Bacteria, Animal Control Slides are for the positive histochemical staining of gram positive and gram negative bacteria in a naturally occurring infection.

 

NEWCOMER SUPPLY VALIDATION PROCEDURE:

  1. Heat dry sections in oven according to your laboratory protocol.
  2. 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 #1 and #2.
  3. Stain in Solution A: Crystal Violet Stain 1%, Aqueous, Brown-Hopps for 2 minutes.
  4. Rinse well in distilled water, ensuring excess stain is removed.
  5. Mordant in Solution B: Iodine, Gram, Aqueous for 2 minutes.
    1. Sections should turn black.
  6. Rinse well in running tap water, ensuring excess iodine is removed.
  7. Blot excess water from slide; decolorize one slide at a time in Acetone, ACS (Part 10014) until blue color stops running; 1-2 dips.
    1. Sections should be very light gray in color.
  8. Quickly rinse in running tap water to remove excess Acetone.
  9. Stain in Solution C: Basic Fuchsin Stain 0.25%, Aqueous; 5 minutes.
  10. Rinse well in running tap water.
  11. Differentiate in Solution D: Gallego Solution for 5 minutes.
  12. Rinse in running tap water.  Blot water off slide(s), but not to dryness.
    1. Proceed with Steps #13 to #16 one slide at a time.
  13. Dip quickly in Acetone, ACS (Part 10014); 1-2 dips.
  14. Dip directly in Solution E: Picric Acid-Acetone 0.05%; 3-10 dips.
  15. Dip quickly in Solution F: Acetone-Xylene 1:1; 5 dips.
  16. Clear in three changes of xylene, 10 dips each; coverslip with  compatible mounting medium.

 

RESULTS:

Gram positive bacteria Blue/violet
Gram negative bacteria Red
Nonreactive lung Negative for gram positive/negative bacteria

 

PROCEDURE NOTES:

  1. Drain slides after each step to prevent solution carry over.
  2. Do not allow sections to dry out at any point during procedure.
  3. If using a xylene substitute, closely follow the manufacturer’s recommendations for deparaffinization and clearing steps.

 

REFERENCES:

  1. Brown, Robert C., and Howard C. Hopps. “Staining of Bacteria in Tissue Sections: A Reliable Gram Stain Method.” American Journal of Clinical Pathology 60.2 (1973): 234-240.
  2. Carson, Freida L., and Christa Hladik Cappellano. Histotechnology: A Self-instructional Text. 4th ed. Chicago: ASCP Press, 2015. 222-224.
  3. Modifications developed by Newcomer Supply Laboratory.