Asparagine Proline Broth

Asparagine Proline Broth

Chemically Defined Medium for Rapid-Growing Mycobacteria
Catalog Number: AS-1126

Overview

Asparagine Proline Broth is a chemically defined, minimal liquid medium used as a key phenotypic test for the identification of the Mycobacterium fortuitum–chelonae complex, including Mycobacterium fortuitum, M. chelonae, M. abscessus, M. peregrinum, and M. mucogenicum.

Only members of this rapid-growing mycobacterial group are able to utilize L-asparagine as the sole nitrogen source and L-proline as the sole carbon source, resulting in visible turbidity or sediment formation within 3–7 days of incubation. Slow-growing mycobacteria and most other non-tuberculous mycobacteria do not grow in this medium.

This test remains recommended in reference laboratory workflows and by international guidelines when molecular methods are unavailable or as a phenotypic confirmation step.

Applications

• Rapid presumptive identification of the Mycobacterium fortuitum–chelonae complex

• Differentiation of rapid-growing mycobacteria from Mycobacterium tuberculosis complex and other NTM

• Component of classical phenotypic test batteries for rapid growers
  (e.g. 5 % NaCl tolerance, arylsulfatase, iron uptake)

Key Features & Benefits

• Highly specific growth response limited to the M. fortuitum–chelonae group

• Chemically defined formulation with minimal background interference

• Clear baseline medium allows easy visual detection of turbidity

• Positive reactions typically observed within 3–5 days at 30 °C

Principle of the Medium

Asparagine Proline Broth contains L-asparagine as the sole nitrogen source and L-proline as the sole carbon source. Rapid-growing mycobacteria of the M. fortuitum–chelonae complex are able to metabolize both substrates efficiently, resulting in visible growth. Organisms lacking this metabolic capability remain in a clear, growth-free medium.

Composition (per litre)

• L-Asparagine – 3.0 g

• L-Proline – 5.0 g

• Potassium dihydrogen phosphate (KH₂PO₄) – 1.0 g

• Disodium hydrogen phosphate (Na₂HPO₄) – 1.0 g

• Magnesium sulfate heptahydrate (MgSO₄·7H₂O) – 0.5 g

• Calcium chloride dihydrate (CaCl₂·2H₂O) – 0.001 g

• Zinc sulfate heptahydrate (ZnSO₄·7H₂O) – 0.001 g

• Copper sulfate pentahydrate (CuSO₄·5H₂O) – 0.001 g

• Ferrous sulfate heptahydrate (FeSO₄·7H₂O) – 0.001 g

Final pH: 6.8 ± 0.2 at 25 °C

Preparation

• Dissolve all inorganic salts in 900 mL distilled water

• Add L-asparagine and L-proline; warm gently if required

• Adjust pH to 6.8

• Make up to 1 L with distilled water

• Filter-sterilise through a 0.22 µm membrane
  (do not autoclave; amino acids are heat-sensitive)

• Dispense 3–4 mL aseptically into sterile screw-cap tubes

Inoculation & Incubation

• Inoculate with 1–2 loopfuls of fresh growth from Löwenstein–Jensen or Middlebrook 7H11 agar

• Include an uninoculated control tube

• Incubate at 30 ± 2 °C with caps loosened

• Examine daily for up to 7 days
  (growth may be visible as early as 3 days)

Interpretation

• Positive: Distinct turbidity or heavy sediment formation
  (indicative of M. fortuitum–chelonae complex)

• Negative: Medium remains crystal clear

Storage & Stability

• Filter-sterilised broth: Store at 2–8 °C, protected from light

• Shelf life: Use within 3 months

Intended Use

For laboratory identification and research use only.
Not for direct clinical diagnostic use.

Quality & Compliance

• Reference phenotypic medium for rapid-growing mycobacteria

• Manufactured under controlled conditions for batch consistency

• Suitable for research, teaching, and reference laboratories

Customs & Trade Information

HS / AHECC Code: 3821.00.00
Prepared culture media for development or maintenance of microorganisms