TECHNICAL DATA SHEET #460 REV.

LOEFFLER MEDIA
PRODUCT:
Tubed Media:
Loeffler Media T6820

PURPOSE:
Loeffler Media is primarily used for the isolation of Corynebacterium diphtheriae from clinical specimens. It is also used to determine the proteolytic activity and pigment production of various microorganisms.

PRINCIPLE:
The present formula for Loeffler Media is a modification of the formula first developed by Loeffler in 18874. Beef extract, dextrose,
proteose peptone, and beef serum provide the nutrients necessary for the growth of the fastidious microorganism,
C. diphtheriae. It is especially useful when small numbers of the microorganisms are present in clinical specimens.1 Recovery is
further enhanced when Loeffler Media is used in combination with Potassium Tellurite Cystine Agar Media and blood agar. On
Loeffler Media, C. diphtheriae produces characteristic metachromatic granules that can be detected by staining with methylene
blue; it also demonstrates the characteristic Chinese-letter formation which is due to the cells snapping off when they divide.
With the above information, a presumptive identification can result in 16-18 hours.
Loeffler Media enhances the demonstration of the organisms natural morphological and physiological characteristics, i.e. pigmentation. It can restore properties after prolonged subculturing and due to the serum content in the media, proteolytic activity,
the enzymatic hydrolysis of proteins into proteoses and peptones, can be demonstrated.

FORMULA:
Approximate, per 250 ml of deionized filtered water.
Loeffler Media:
Beef Extract ......................................................0.75 g
Dextrose ...........................................................1.25
Sodium Chloride ...............................................1.25
Proteose Micro Peptone ...................................2.50
Horse Serum ................................................750.00 ml
Final pH 7.1 0.2 at 25C

PRECAUTIONS:*
For in vitro diagnostic use. Observe approved biohazard precautions.
Storage: Upon receipt store at 2-8C away from direct light. Media should not be used if there are signs of contamination,
deterioration (cracking, shrinking, or discoloration), or if the expiration date has passed.
Limitations: Microscopic morphology may vary with different batches/lots of Loeffler Media.
C. diphtheriae type gravis is the least likely to be recognized on microscopic examination.3
Nasopharyngeal and throat specimens should both be obtained when C. diphtheriae is suspected since 20% of positive cultures
can be missed when only one site is cultured.3
Recovery of C. diphtheriae requires culturing on selective and nonselective media.

PROCEDURE:*
Specimen collection: Information on specimen collection is found in standard reference material. In general, specimens should
be protected from extreme heat and cold and should be delivered to the laboratory without delay. Nasopharyngeal cultures
should be obtained using an alginate swab that reaches deep into the posterior nares. For throat cultures rub vigorously over
any inflammatory lesion. Specimens can be directly plated onto a Loeffler slant if transport time is less than 24 hours.
Method of Use: Prior to inoculation, the media should be brought to room temperature. Remove excess fluid at the base of the
slant. Inoculate the slant with the specimen swab in a fishtail motion and incubate aerobically for 8-24 hours at 35C. Prepare
smears and stain with methylene blue stain. Colonies showing metachromatic granules and Chinese-letter formation on methy-

27120 SW 95th Avenue Wilsonville, OR 97070 800.547.0659

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TECHNICAL DATA SHEET #460 REV. 3

lene blue stain are definitively identified through biochemical testings and by tests for toxigenicity.
To detect proteolytic ability, inoculate aerobic microorganisms directly on the surface of the Loeffler slant; with anaerobes incubate anaerobically or fill the slant with thioglycollate broth before incubation. Incubate aerobically at 35C for 3-4 days, possibly
longer depending on the microorganism. Examine for colonies surrounded by small craters of liquefied media or liquefication of
the slant with the production of a putrefied odor.
Interpretation:3
Corynebacterium diphtheriae Biotypes:

Metachromatic Granules:

Morphology:

mitis

Intensely stained

Long, pleomorphic but rigid

rods

intermedius

Cells frequently swollen

with terminal granules
Irregularly barred

Highly pleomorphic
with very long and
short rods

gravis

No granules but alternate

bands of light and dark blue

Short rods with many

cocci of pear-shaped forms

Material Required but Not Provided: Standard microbiological supplies and equipment commonly found in a microbiological
laboratory are not provided.

QUALITY CONTROL:*
Microorganisms Used (ATCC #):
Corynebacterium diphtheriae (13812)
Corynebacterium pseudodiphtheriticum (10700)

Expected Results:
Growth
Growth

User Quality Control: Check for signs of contamination and deterioration. Loeffler Media should appear moderately firm,
opaque, and white in color.

BIBLIOGRAPHY:
1.
2.
3.
4.

Forbes, B. A., et al., Bailey and Scotts Diagnostic Microbiology, 12th ed., C. V. Mosby, St. Louis, 2007.
Koneman, E. W., et al., Color Atlas and Textbook of Diagnostic Microbiology, 6th ed., J. B. Lippincott, Philadelphia, 2005.
Murray, P.R., et al., Manual of Clinical Microbiology, 9th ed., American Society for Microbiology, Washington, D. C., 2007.
Loeffler, F., Centr. f. Bakt., 2:105, 1887.

* For more detailed information, consult appropriate references.

PML Microbiologicals, Inc.
Data #460
Copyright 1989 by PML Microbiologicals, Inc.
Revision Date: October 2008

27120 SW 95th Avenue Wilsonville, OR 97070 800.547.0659

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