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Microbial limit test (MLT) – Testing Procedure

The Microbial limit test (MLT) is performed to assess how many and which of certain viable microorganisms are present in non-sterile pharmaceutical, healthcare or cosmetics manufacturing samples that range from raw materials to finished products.

Microbial limit test (MLT)

1.0   Objective :

    • To lay down a procedure for microbial limit test (MLT).

2.0   Scope :

    • This SOP is applicable in microbiology for estimation of microbial contamination by performing Microbial Limit Test (MLT) in Raw Materials, Finished Products.   

3.0   Procedure – Microbial Limit Test (MLT) :

    • Sample Preparation for Microbial Limit Test (MLT):

    • Weigh and transfer 10 ml for liquid/ 10 g. for solid of sample, in Test tube/ Flask (Solution A) containing 90 ml / 100 ml of sterile pre incubated Soya bean Casein Digest Medium (SCDM) or buffered Sodium Chloride Peptone Solution (if required adjust pH to 7.0).
    • Prepare the medium.
    • If the sample, to be examined, has antimicrobial activity this may be adequately neutralized by adding neutralizing agents to SCDM or buffered Sodium Chloride-Peptone Solution.
    • The given below neutralizers may be added.
    • Neutralizing agents may be used to neutralize the activity of antimicrobial agents.
    • They may be added to the chosen diluent or the medium preferably before sterilization.
    • If used, their efficacy and their absence of toxicity for micro-organisms must be demonstrated by carrying out a blank with neutralizer and without product.
Type of antimicrobial agent In activator Concentration Remarks
Phenolics, Parahydroxy benzoate (parabens) Polysorbate 80 30 g per litre Add after Sterilization of the diluents
iodine, Quaternary Ammonium Compounds (QACs), Lecithin

Sodium lauryl sulphate

 3 g per litre

4 g per litre

 Add after Sterilization of the diluents
Alcohol, Aldehydes, sorbates Dilution ………… after Sterilization of the diluents
Mercurials, halogens Sodium Thiosulphate 5 g per litre Add prior to Sterilization

    • Mix well to obtain a uniform solution / suspension of required dilution as described below.

Dilution ratio Product quantity Quantity of diluents
1 : 10 10 ml / 10 g 90 ml / 100 ml
1 : 20 5 ml / 5 g 95 ml /100 ml
1 : 50 1 ml / 1 g 49 ml / 50 ml
1 : 100 1 ml / 1 g 99 ml/ 100 ml

  • Enumeration of total viable aerobic count – Microbial Limit Test (MLT):

  • By Pour-plate method:

    • Total Aerobic Microbial Count (TAMC):

    • Add 1 ml of the final dilution (Solution A) to each Petri dish than add approximately 15 to 20ml of sterile Soyabean Casein Digest Agar, in to two Sterile Petri dishes of 90mm and mix the contents of Sterile Petri dishes by rotating and tilting the plate, and allow medium to solidify.
    • The temperature of media during transfer should be approximately 45°C. Incubate these Sterile Petri dishes at 30 to 35°C for 3 to 5 days. Keep the plates in inverted position.

    • Total Combined Yeasts and Mold Count (TYMC):

    • Add 1 ml of the final dilution (Solution A) to each Petri dish.dd approximately 15 to 20ml of sterile Sabouraud Dextrose Agar, in two Sterile Petri dishes of 90mm.
    • Mix the contents of Sterile Petri dishes by rotating and tilting the plate, and allow medium to solidify.
    • The temperature of media during transfer should be approximately 45°C.
    • Incubate these Sterile Petri dishes at 20 to 25°C for 5 to 7 days.
    • Keep the plates in inverted position.
    • After completion of incubation period, take the arithmetic mean of the count per medium, and calculate the number of CFU per g. or ml of the product.

    • Negative Control:

    • To verify testing conditions, a negative control shall be performed using the chosen diluents in place of the test preparation.
    • There must be no growth of micro-organisms.

    • Positive Control:

    • Inoculate portions/plates of Soya bean Casein Digest Agar / Sabouraud-dextrose agar with a small number (not more than 100 CFU) of the micro-organisms.
    • Incubate in the conditions as stated above.
    • Interpretation of results:
    • In general the following calculation can be used

    Average Plate count                                    1

        =—————————————-   x  ———————– =…………….cfu/g or ml

Volume of test Article/sample              Dilution factor

  • Testing & Identification of Products (Specific Micro-organism):

    • Escherichia coli – Microbial Limit Test (MLT):

    • Sample preparation and pre-incubation:
    • Transfer 10 ml of Solution A in 90 ml pre incubated Soya Bean Casein Digest Medium (SOLUTION-B)., mix and incubate at 30 – 35°C for 18 – 24 hours.
    • Selection and subculture:
    • Shake the container, transfer 1 ml of SOLUTION-B to 100 ml of pre incubated MacConkey broth and incubate at 42 – 44°C for 24 – 48 hours.
    • After incubation period subculture it on a plate of pre incubated MacConkey agar and incubate at 30 – 35 °C for 18 – 72 hours.
    • Interpretation:
    • The growth of colonies indicates the possible presence of Escherichia coli.
    • This will be confirmed by identification test.
    • The product complies with the test if no colonies are present or if the identification tests are negative.
Medium Description of colony
MacConkey agar Brick red may have surrounded zone of precipitated bile.

    • Positive Control:

    • Use culture of Escherichia coli ATCC8739/MTCC1687.
    • Negative Control:
    • Carry out the negative control by using one set of tube / Petri plates of sterile culture media.
    • Identification (Indole test):
    • Incubate the suspected colonies into tube containing 10 ml of 0.1 % peptone water and incubate the tube at 30°C to 35°C for 18 – 24 hours.
    • After completion of incubation add 0.5 ml of Kovac’s reagent, shake well and allow standing for one minute.
    • The appearance of cherry red colour ring along the side of the test tube confirms the presence of E. coli.
    • The product complies the test if the confirmatory identification tests are negative.

    • Staphylococcus aureus – Microbial Limit Test (MLT):

    • Selection and subculture;-
    • Take a loop full culture from SOLUTION-B and subculture on to the surface of Pre incubated Mannitol Salt Agar Plate and incubate at 30 – 35°C for 18 to 72 hours.
    • Interpretation:
    • The possible presence of S. aureus is indicated by the growth of yellow or white colonies surrounded by yellow zone.
    • This will be confirmed by identification test.
    • The product complies with the test if no colonies are present or if the identification tests are negative.
    • Positive Control: Use culture of Staphylococcus aureus ATCC6538/MTCC737.
    • Negative Control: Carry out the negative control by using one set of tube / Petri plates of 90mm sterile culture media.

    • Identification (Coagulase test):

    • With the aid of an inoculating loop, individually transfer suspected colonies to separate tubes containing 0.5 ml of mammalian plasma (preferably rabbit or horse).
    • Incubate in a water-bath at 37°C±1°C for 3 to 24 hours, in parallel with positive control using known strain of Staphylococcus aureus and negative control using Plasma alone.
    • Examine after 3 hours and at suitable intervals for 24 hours thereafter for the presence of coagulation.
    • If coagulation in any degree is observed, the presence of Staphylococcus aureus is indicated.
    • Record the results.
    • The product complies with the test if colonies of types describes are not present or if the confirmatory identification tests are negative.

Test for Staphylococcus aureus

Selective medium Characteristic colonial morphology Gram stain
Vogel-Johnson agar Black surrounded by yellow zones Positive cocci
( in clusters)
Mannitol-salt agar Yellow colonies with yellow zones Positive cocci
( in clusters)
Baird-Parker agar Black, shiny, surrounded by clear zones of 2 to 5 mm Positive cocci
( in clusters)

  • Pseudomonas aeruginosa – Microbial Limit Test (MLT):

    • Selection and subculture:
    • Subculture a portion from SOLUTION-B on to the surface of Pre incubated Cetrimide agar Plate and incubate at 30 – 35°C for 18 to 72 hours.
    • Take a loop full culture from Cetrimide agar plate and subculture on Pre incubated pseudomonas flouresein agar for the detection of Pyocyanin and incubate at 33.0°C to 37.0°C for not less than 72 hrs.
    • After completion of incubation examine under UV light.
    • Interpretation:
    • The growth of colonies indicates the possible presence of P. aeruginosa.
    • This will be confirmed by identification test.
    • The product complies with the test if no colonies are present or if the identification tests are negative.

    • Positive Control:

    • Use culture of Pseudomonas aeruginosa ATCC9027/MTCC1688.
    • Negative Control:
    • Carry out the negative control by using one set of tube / petri plates of sterile culture media.
    • Identification (Oxidase test):
    • With the aid of an inoculating loop, transfer suspected colonies to strip or discs of filter paper impregnated with N, N-dimethyl-p-phenylenediamine dihydrochloride
    • If a purple colour produced within five to ten seconds conforms the presence of P.
    • Record the result.
    • The product complies with the test if the confirmatory identification tests are negative.

Test for Pseudomonas aeruginosa

Medium Characteristic colonial morphology Fluorescence in UV light Oxidase test Gram stain
Cetrimide agar Generally greenish Greenish Positive Negative rods
Pseudomonas agar medium for detection of fluorescein Generally colorless to yellowish Yellowish Positive Negative rods
Pseudomonas agar medium for detection of Pyocyanin Generally greenish Blue Positive Negative rods

  • Salmonella ssp – Microbial Limit Test (MLT):

    • Preparation of Sample:
    • Weigh and transfer 10 ml / 10 g. of sample, in test tube containing 90 ml / 100 ml of Pre incubated soya bean casein digest medium, mix and incubate at 30°C to 35°C for 18 to 24 hrs (SOLUTION-C).
    • Selection and Subculture:
    • Transfer 0.1 ml of soya bean casein digest medium to 10 ml of pre incubated Rappaport Vassiliadis Salmonella enrichment broth and incubates at 30 – 35 °C for 18 – 24 hours.
    • Subculture on pre incubated plates of xylose, lysine, deoxycholate agar. Incubate at 30 – 35 °C for 18 – 48 hours.
    • Interpretation: The possible presence of Salmonella is indicated by the growth of well-developed, red colonies, with or without black centers.
    • This will be confirmed by identification test.
    • The product complies with the test if no colonies are present or if the identification tests are negative.

    • Positive Control:

    • Use culture of Salmonella NCTC6017/MTCC3858.
    • Negative Control:
    • Carry out the negative control by using one set of tube / Petri plates of 90ml of sterile culture media.
    • Identification (Triple sugar iron agar test):
    • Individually transfer the suspected  colony  by first  sub culturing  the slope of  slant of  Triple Sugar-Iron  Agar  with inoculating loop .
    • Stabbing with inoculating straight wire well in the butt. Incubate at 30°C to35°C for 18 to 24 hours.
    • After incubation, examine the tube of Triple Sugar Iron Agar Medium for the presence of microbial growth    and    for   the   following    Physical characteristics:
      • Color changes from red slant to yellow butt, with or without concomitant blackening of butt due to production of H2S in agar.
      • If the butt, slant of Triple Sugar Iron Agar shows growth and physical characteristics confirming to the above descriptions the presence of Salmonella species is indicated.
    • Record the observations.
    • The product complies with the test if the confirmatory identification tests are negative.

Test for Salmonella

Medium Description of colony
Bismuth sulphite agar Black or green
Brilliant green agar Small, transparent and colorless, or opaque, pinkish or white (frequently surrounded by a pink or red zone)
Deoxycholate-citrate agar Colorless and opaque, with or without black centers
Xylose-lysine-desoxycholate agar Red with or without black centers

    • Test for Bile-tolerant gram-negative bacteria:

Note: Use the media of SOLUTION- A for test of Bile-tolerant gram-negative bacteria.

    • Use part sample of tube -A incubate at 20-25°C for 2 to 5 hours.
    • After incubation transfer the quantity from this tube-A corresponding to 1 g or 1ml of the product to 100 ml Pre incubated Enterobacteria Enrichment Broth Mossel and incubate at 30°C to 35°C for 24 to 48 hours, subculture on Pre incubated Violet red bile glucose agar plate, incubate at 30°C to 35°C for 18-24 hours,
    • The product complies with the test if there is no growth of colonies.
    • If there is growth then proceeds further for Quantitative test.
    • Mix Enterobacteria Enrichment Broth Mossel after incubation and inoculate suitable quantities of EE broth containing respectively 0.1 g, 0.01 g and 0.001 g (or 0.1ml, 0.01 ml, 0.00 1ml) of the product to be examined.
    • Incubate at 30°C to 35°C for 24 to 48 hours.
    • Subculture each of the cultures on a pre incubated plate of Violet red bile glucose agar to obtain selective isolation.
    • Incubate the plates at 30°C to 35°C for 18 to 24 hours.
    • Growth of colonies constitutes a positive result.
    • Note the minimum quantity of product which gives positive result and maximum quantity of the product that gives a negative result.
    • Determine the probable number of bacteria from table given below and record the result in Annexure-1.

    • Result for each quantity of product.

0.1 gm or 0.1 ml 0.01 gm or 0.01 ml 0.001 gm or 0.001 ml Probable number of bacteria per gram of product.
+ + + More than 1000
+ + – Less than 1000 and More than 100
+ – – Less than 100 and More than 10
– – – below 10

    • Test for Clostridium species – Microbial Limit Test (MLT):

    • Use part sample of Solution A take two equal portions corresponding to 1 gm or 1 ml of the product to be examined.
    • Heat one portion to 80°C for 10 min and cool rapidly, do not heat the other portion.
    • Selection and Subculture: Transfer the 10 ml of each of the homogenized portions to two tubes containing 100 ml of Pre incubated Reinforced Medium for Clostridia.
    • Incubate under anaerobic condition at 30°C to 35°C for 48 hours. After incubation make sub-culture from each tube on Columbia agar.
    • Incubate under anaerobic conditions at 30°C to 35°C for 48 hours.

    • Interpretation:

    • If no anaerobic growth of microorganisms is detected on Columbia agar, the product complies with the test.
    • If anaerobic growth of microorganisms is present then perform Catalase test as given  
    • Place a drop of hydrogen peroxide on a clean slide and Take a portion of suspected colony & rub with drop of hydrogen peroxide then it will generate effervescences of oxygen bubble.
    • it indicates the positive catalase test
    • The occurrence of anaerobic growth of rods (with or without endospores) giving a negative catalase reaction indicates the presence of clostridia.

    • Test for Candida albicans – Microbial Limit Test (MLT) :

    • Use 10 ml SOLUTION- A or the quantity corresponding to not less than 1g or 1 mL, to inoculate 100 mL of pre incubated Sabouraud Dextrose broth, & mix it properly.
    • Incubate at 30°C to 35°C for 3-5 days.
    • After incubation make sub-culture on a pre incubated plate of Sabouraud-dextrose agar and incubate at 30 – 35°C for 24 to 48 hours.
    • Interpretation:
    • Growth of white colonies may indicate the presence of C. albicans.
    • The product complies with the test if such colonies are not present.
    • Positive Control:
    • Use culture of Candida albicans ATCC 10231/MTCC 227.
    • Negative Control: Carry out the negative control by using one set of tube / Petri plates of 90ml of sterile culture media.
    • If colonies are found, identification shall be performed by identification test as follow.
    • Identification:
    • Place a drop of mounting fluid (Lacto phenol cotton blue) on a clean slide.
    • Transfer a small tuft of the suspected colony into the drop using a flamed, cooled needle.
    • Gently teased the material using two mounted needle & mix gently the stain.
    • Place the cover slip over the preparation taking care to avoid air bubble.
    • Observe under microscope If it shows Pseudomycelial growth & multilateral budding then it indicates the presence of C. albicans
    • The product complies with the test if the confirmatory identification tests are negative.
    • Note:-Send culture suspensions/Suspected colonies outside in a authorized Laboratory or organization for identification if Biochemical test kits are not available.

    • Test for Shigella boydii – Microbial Limit Test :

    • After incubation shake the flask of Solution-C and transfer 1 ml to 100 ml Pre incubated GN Broth and incubate at 30º to 35º C for 24 to 48 hours.
    • Further subculture on a pre incubated plate of Xylose Lysine Deoxycholate Agar and incubates at 30º to 35º C for 24 to 48 hours.
    • If colorless, opaque colony without black centre appears on the medium as mentioned above, indicates possible presence of Shigella.
    • Then proceed for further confirmation tests.
    • If such characteristic growth doesn’t appear then sample passes the test for absence of Record the results in annexure-I.
    • Positive Control:
    • Use culture of Shigella boydii ATCC 12030/MTCC 11947.
    • Negative Control:
    • Carry out the negative control by using one set of tube / Petri plates of 90ml of sterile culture media.
    • Further confirmation can be done by sub culturing the suspected colonies on Modified SS agar.
    • If the characteristic growth than confirms the presence of Shigella in the sample under examination, sample fails the test for absence of Shigella.
    • Investigate the out of specification results to follow handling and investigation of out of specification result in Microbiology testing.
    • In case of any conditional release of Raw material/Finished product, Microbiologist shall release the same on the basis of three Day count of TAMC and TYMC obtained on annexure – III and final release of the Raw material/Finished product shall be done after the completion of complete analysis along with pathogen analysis.
    • On final release of Raw material/Finished product, Microbiologist shall attach the results of three day count of TAMC and TYMC shall be recorded on Annexure – III with final report.

4.0   Glossary :

SCDM Soya bean Casein Digest Medium CFU Colony forming units
TAMC Total Aerobic Microbial Count TYMC Total Combined Yeast and Mold Count
ATCC American Type Culture Collection MTCC Microbial  Type Culture Collection
TSA Triple Sugar Iron Agar NCTC National Culture Type Collection
I.P Indian Pharmacopeia E.P European Pharmacopeia
U.S.P United State Pharmacopoeia GN Gram Négative Broth

5.0   Reference Document:

    • IP 2018: Chapter No. 2.2.9 (Microbiological contamination in non sterile product)
    • USP-NF-41: Chapter No. 62 (Microbiological Examination of non sterile product and test for specified organisms)
    • EP 9.0 : Chapter No.2.6.12 Microbiological examination of non sterile product (Total viable aerobic count)
    • EP 9.0: Chapter No.2.6.13 Microbiological examination of non sterile product (Specified Micro organisms)

4.0   Annexure – Microbial Limit Test:

Annexure I: Microbial limit test (MLT) report

Name of Sample      :                             Date of  Test  :                                                                        Batch No./A.R. No. :                           Date of Report  :

Analyzed as per      :   IP / BP / USP / In-House                   Test performed by :
Sample Preparation (Pretreatment):  Transfer _______ g/ ml of sample in 100 ml/ 90 ml Buffered Sodium Chloride Peptone Solution / Soyabean Casein Digest Broth / Phosphate Buffer Solution (Solution A). Media Lot No._____________________. Mix well to dissolve it completely.
­TOTAL VIABLE AEROBIC COUNT:

Method used      :     Pour plate method.

Procedure          :      Place 1 ml of sample of pretreated sample (Solution-A) equivalent to 1 g or 1 ml of sample.                                
Step CFU Mean  x Dilution factor CFU/ g/ ml Limit
1)  TOTAL AEROBIC MICROBIAL COUNT

Medium:  Soyabean casein Digest Agar, Media Lot No. ______________

Incubation: 30 °C to 35°C for 3 to 5 days Incubator ID No.:                                

1.

2.
Mean:
Organism Used for positive control: Positive Control: Negative Control: 
2)  TOTAL COMBINED YEAST/MOLD COUNT

Medium: Sabouraud dextrose Agar, Media Lot No._________________

Incubation: 20°C to 25°C for 5 to 7 days, Incubator ID No.:                               

1.

2.
Mean :
Organism used for positive control : Positive Control : Negative Control :

B)TEST FOR SPECIFIED ORGANISMS

Step Sign / Date
From Solution A transfer 10 ml of sample in 90 ml sterile Soyabean Casein Digest Medium, Media Lot No. _______________________, Incubated at 30° to 35° C for 18 to 24 hours (Solution-B).
Test Results: Growth Observed / Not Observed Negative Control Positive Control Organism Used  

 
1) Test for Escherichia coli  :                                Sign / Date
Carry out positive control by using culture of Escherichia coli ATCC 8739/MTCC 1687

Carry out Negative control by using one set of tube / Petri plate of sterile culture media.
Transfer 1.0 ml of Solution B to 100 ml of pre incubated MacConkey broth. Media Lot No.____________. Incubate at 42° to 44° C for 24 to 48 hours
Test Results: Growth Observed / Not Observed Positive Control Negative Control
Subcultures on plates of preincubated MacConkey Agar and incubate at 30° to 35° C for 18 to 72 hours.
Media Lot No. Standard Result Test Result Positive Negative Gram Nature
  Brick red may have surrounded zone of precipitated bile. Observed /Not Observed

Identification:
If positive, sub culture suspected colonies on pre incubated EMB agar plate and incubate at 30° to 35° C for 48 hours.
Test Result: Characteristic metallic sheen under reflected light and a blue black appearance under transmitted light: Observed/ Not observed.
Positive Control = Negative Control = Gram Nature =
If positive, inoculate suspected colonies into tube containing 10 ml of 0.1% peptone water. Incubate at 30° to 35° C for 18 to 24 hours. After completion of incubation add 0.5 ml of Kovacs reagent, shake well, and allow standing for one minute.
Test Result:  Appearance of cherry red colour ring: Observed / Not observed. Positive Control =
Conclusion: Escherichia coli   Present / Absent per _____ g /ml  of sample.

2)     Test for Staphylococcus aureus  :  

Sign / Date
Carry out positive control by using culture of Staphylococcus aureus ATCC 6538/MTCC 737

Carry out Negative control by using one set of tube / Petri plate of sterile culture media.
Take a loop full culture a portion from Solution B and sub culture on the surface of pre incubated Mannitol Salt agar plate (MSA). Incubate Mannitol Salt agar Plates at 30˚C to 35˚C for 18 to 72 hours.
Media Lot No. Standard Results Test Results Positive Negative Gram Nature
  Yellow colonies with yellow zones. Observed / Not Observed
Identification: If growth of suspected colonies occurs carry out Coagulase test.

Transfer suspected colonies using inoculating loop in to tubes containing 0.5 ml of mammalian plasma. Incubate at 37 ± 1˚ C for 3 to 24 hours in water bath. Examine the tubes at 3 hours and subsequently at suitable interval for 24 hours.
Test Results; Coagulation Observed / Not Observed.
Conclusion: Staphylococcus aureus Present / Absent per _____ g /ml of sample.

2)     Test for Pseudomonas  aeruginosa  :  

Sign / Date
Carry out positive control by using culture of Pseudomonas aeruginosa  ATCC 9027/MTCC 1688

Carry out Negative control by using one set of tube / Petri plate of sterile culture media.
Sub culture a portion from Solution B on a Pre incubated Cetrimide Agar plate, Incubate at 30o to 35oC for 18 to 72 hours.
Media Lot No. Standard Result Test Result Positive Negative Gram Nature
  Greenish colonies Observed / Not Observed
Identification:                       
Sub culture suspected colony on of pre incubated Pseudomonas agar plate for the detection of Fluorescein and pre incubated Pseudomonas agar plate for the detection of Pyocyanin.

Incubate at 33° to 37° C for not less than 72 Hrs. After completion examine under UV light.
Result:

1. Pseudomonas agar medium for the detection of Fluorescein: Yellowish Fluorescence. Gram Negative   Observed/ Not Observed.

2. Pseudomonas agar medium for the detection of Pyocyanin: Blue Fluorescence. Gram Negative.           Observed/ Not Observed.
If growth of suspected colonies occurs carry out Oxidase test.

Transfer suspected colonies using inoculating loop on to Oxidase test disc [N-N-dimethyl- p-phenylene diamine oxalate].
Result : Development of purple colour in 5 to 10 seconds:                      Positive/ Negative
Conclusion: Pseudomonas aeruginosa Present / Absent per ­­­­­­­­­­­­­­­­­­­­­­­­­­­­________ g /ml of sample.

Annexure II: Microbiological Examination of Raw Materials & Finished Products

Item / Product Name: …………………………………………………………..……………………………

Batch No.:………………….…………                                          A.R. No. : ……………………………

Mfg. Date: …………………………….                                        Exp. Date: …………………………..

Date of Sampling            : ………………….                          Sampled Quantity:…………………

Sampled By: ………………….…                                              Date of analysis: ………………….…

Completion Date: ………………….

 Test  Media Lot No. Observation Acceptance Criteria
Total Aerobic Microbial Count
Total combined Yeasts & Mold count
E. Coli
Staphylococcus aureus
Pseudomonas aeruginosa
Clostridia sporogenes
Candida albicans
Salmonella
Enterobacteriacae
Shigella boydii

Annexure III: Conditional Release of Microbial Analysis of Raw Material/ Finished Product

      Item / Product Name:.…………………………………………………………..…………………….

      Batch No.:                   ………………….…….                                     A.R. No. :     ……………………….……

      Mfg. Date:                    ………………….……                                     Exp. Date:     ……………………………

      Date of incubation      ………………….…….                               Time of incubation: ……………..……

      Date of review of result ………………………                          Time of review of result ………………

 Test  Media Lot No.  Observation Mean  x Dilution factor Acceptance Criteria
Total Aerobic Microbial Count 1.

2.
Mean:
Organism Used for +ve control: Positive control: Negative Control:
Total Yeast & Mold Count 1.

2
Mean:
Organism Used for +ve control:  Positive control:     Negative Control:  

 

 Remarks: Data reviewed and found/Not found well within the specification limit and analysis will be completed on  _____________.

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Janki Singh

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