Applied Microbiology/Pharmaceutical Microbiology and Cleanroom Technology PDF

Applied Microbiology/Pharmaceutical Microbiology and Cleanroom Technology Gillian Carney [email protected] Room; 1163 Cleanroom Technology - Overview - History and Bioprocessing - Cleanroom Design and Contamination - Cleaning methods - Resistance to Disinfectants - Clean in Place - Monitoring...

Applied Microbiology/Pharmaceutical Microbiology and Cleanroom Technology Gillian Carney [email protected] Room; 1163 Cleanroom Technology - Overview - History and Bioprocessing - Cleanroom Design and Contamination - Cleaning methods - Resistance to Disinfectants - Clean in Place - Monitoring in Cleanrooms - Endotoxins and LAL Assay - Operation of Cleanrooms Activity - Based on your understanding of microbial contamination and air pollution summarise the main concerns for a cleanroom environment and how you might control it? Concern Control Measure - Where do you think cleanroom technology is applied? Why is the technology important in these settings.  Activity - Where do you think cleanroom technology is applied? Why is the technology important in these settings. Industry Why CRT is important here Cleanroom Technology What is a Cleanroom? Defined in the International Organization for Standarization (ISO) standard 14644-1 as: A room in which the concentration of airborne particles is controlled, and which is constructed and used in a manner to minimise the introduction, generation, and retention of particles inside the room and in which other relevant parameters, e.g. temperature, humidity, and pressure, are controlled as necessary. The first two thirds of the definition is, in essence, what a cleanroom is. It is a room that minimises the introduction, generation and retention of particles. Cleanroom Technology A room in which the concentration of airborne particles is controlled, and which is constructed and used in a manner to minimise the introduction, generation, and retention of particles inside the room and in which other relevant parameters, e.g. temperature, humidity, and pressure, are controlled as necessary. 1. Air supplied is tightly controlled with high-efficiency filters (also regulate things like temperature, humidity etc). 2. The materials used do not generate particles and can be easily cleaned. Cleanroom Technology This is achieved; Firstly, by supplying it with exceptionally large quantities of air that has been filtered with high efficiency filters. This air is used to (1) dilute and remove the particles and bacteria dispersed from personnel and machinery within the room and, (2) to pressurise the room and ensure that no dirty air flows into the cleanroom. Secondly, a cleanroom is built with materials that do not generate particles and can be easily cleaned. Finally, cleanroom personnel use clothing that envelops them and minimises their dispersion of particles and microorganisms. Cleanrooms can also control the temperature, humidity, sound, lighting, and vibration.  History of Cleanrooms- Cleanrooms in hospitals. 1860s-1900s - Lord Lister *Bacteria were the cause of - 1889 Aberdeen Royal Infirmary - 1890s Royal Infirmary Edinburgh surgical wound infection - Listers spray – carbolic acids in the - Gowns introduced - Antiseptic solution air of the operating room (Carbolic acid) History of Cleanrooms- Cleanrooms in hospitals. 1860s-1900s Listers techniques were further developed into the field of aseptic techniques. Instruments and bandages were boiled Increase in hand washing By 1900, surgical gloves, masks and gowns were introduced. (steam sterilized before operation) 1907 Royal Infirmary, Edinburgh History of Cleanrooms- Cleanrooms in hospitals. 1860s-1900s 1940s – Ventilation was introduced in temperate climates World War II - Ventilation key - Major problem in the crowded situations that occurred e.g submarines, air raid shelters etc. - Airborne samplers were used to study the dynamics of contamination and by 1960s most of the key principles of this technology had been established. - This period also highlighted how people are a major source of infection History of Cleanrooms- Cleanrooms in hospitals. 1860s-1900s Charnley & Howort– Hip replacement 10% sepsis rate in first round of operations Restricted a small area & controlled the air flow Infection rate went to 0.5 um per m3. IS0 Standard 14644-1 A series of cleanroom standards. -Design -Testing -Operation -Biocontamination. The first document, published in 1999, is IS0 14644-1 and entitled ‘Classification of Air Cleanliness’. Now a system based on ISO-14698 documentation  Comparison of FED and ISO Standards maximum particles/m3 a FED STD 209E Class ≥0.1 µm ≥0.2 µm ≥0.3 µm ≥0.5 µm ≥1 µm ≥5 µm equivalent ISO 1 10b d d d d e ISO 2 100 24b 10b d d e ISO 3 1,000 237 102 35b d e Class 1 ISO 4 10,000 2,370 1,020 352 83b e Class 10 ISO 5 100,000 23,700 10,200 3,520 832 d,e,f Class 100 ISO 6 1,000,000 237,000 102,000 35,200 8,320 293 Class 1,000 ISO 7 c c c 352,000 83,200 2,930 Class 10,000 ISO 8 c c c 3,520,000 832,000 29,300 Class 100,000 ISO 9 c c c 35,200,000 8,320,000 293,000 Room air a All concentrations in the table are cumulative, e.g. for ISO Class 5, the 10 200 particles shown at 0,3 μm include all particles equal to and greater than this size. b These concentrations will lead to large air sample volumes for classification. Sequential sampling procedure may be applied; see Annex D. c Concentration limits are not applicable in this region of the table due to very high particle concentration. d Sampling and statistical limitations for particles in low concentrations make classification inappropriate. e Sample collection limitations for both particles in low concentrations and sizes greater than 1 μm make classification at this particle size inappropriate, due to potential particle losses in the sampling system. f In order to specify this particle size in association with ISO Class 5, the macroparticle descriptor M may be adapted and used in conjunction with at least one other particle size. (See C.7.) g This class is only applicable for the in-operation state. Occupancy states inform classification GMP EU classification 1. As built: maximum particles/m3 Installation is complete, services Class At Rest At Rest In Operation In Operation connected and functioning, but with no production equipment, materials or 0.5µm 5µm 0.5µm 5µm personnel present Grade A 3,520 20 3,520 20 Grade B 3,520 29 352,000 2,900 2. At-rest: As built + equipment installed and Grade C 352,000 2,900 3,520,000 29,000 operational in a manner agreed between the Grade D 3,520,000 29,000 Not defined Not defined customer and supplier, but with no personnel present. EU GMP guidelines require cleanrooms to meet particle counts during operation and at rest 3. In Operation: At rest + with the specified number of personnel present and working in the manner agreed upon. Monitoring of Microbial Limits Microbial limits are also monitored and appropriate Alert and Action limits should be set for each cleanroom environment, Grade Glove print Settle plates Contact plates Air Sample 5 fingers on (Diam. 90mm (Diam. 55mm cfu/m3 both hands cfu/4hours) cfu/plate) cfu/glove Grade A 1 1 1 1 Grade B 10 5 5 5 Grade C 100 50 25 - Grade D 200 100 50 - Bioprocessing Bioprocessing uses organisms or biologically derived macromolecules to carry out enzymatic reactions or to manufacture products. Biopharmaceutical; “Biological” “medical product” Defined as complex molecules derived from a biological source, with the purpose to diagnose, prevent, treat, or cure diseases or conditions of human beings. e.g. proteins, vaccines, cell therapies etc Bioprocessing e.g. Production of Insulin Insulin is used to treat Type 1 diabetes. (31,000 new cases per year, EU) Previously treated using insulin isolated from Porcine and bovine pancreatic tissue Development of Recombinant DNA technology allowed for the production of human derived insulin using microorganisms.  Primary Upstream Downstream Formulation Capture & processing processing & Filling Recovery Centrifugation – Challenging step as Concentration separating cells target molecule may from media only be