The following principles should be followed when monitoring the disinfection effect in the hospital: the monitoring personnel should undergo professional training, master certain disinfection knowledge and have skilled inspection skills; Choose a reasonable sampling time (after disinfection before use); Follow strict aseptic operation.
20.2 Monitoring method of thermal sterilization effect
20.2. 1 pressure steam sterilization effect monitoring method
20.2. 1 chemical monitoring method
(1) monitoring method of chemical indicator card (tube): put the chemical indicator tube (card) which can indicate both temperature and temperature duration in the center of each package to be sterilized, take out the indicator tube (card) after a sterilization cycle, and judge whether the sterilization conditions are met according to the changes of its color and properties.
(2) Chemical indicator tape monitoring method: stick the chemical indicator tape outside each package to be sterilized, and observe the color change after a sterilization cycle to indicate whether it has been sterilized.
(3) Result judgment: When the indicator tube (card) is tested, if the character or color of the indicator tube (card) changes to the specified conditions, it can be considered that the packaging is qualified for sterilization.
(4) Precautions: Chemical indicators used for monitoring must be approved by the Ministry of Health and used within the validity period.
20.2. 1.2 biological monitoring method
(1) indicator strain: the indicator strain is Bacillus stearothermophilus spore (ATCC7953 or SSIK3 1 strain), and the bacterial content of the bacterial slice is 5.0x105-5.0x106cfu/slice, at12/.
(2) Culture medium: The experimental culture medium is bromocresol purple peptone water culture medium.
(3) Detection method: Put two pieces of Bacillus stearothermophilus into a small sterile paper bag for human use and put it in the center of the standard test package.
Place the standard test kit (3 ordinary long-sleeved surgical gowns, 4 small surgical towels, 2 Chinese surgical towels, 1 large surgical towel, 30 pieces 10cm x 10cm8 gauze wrapped 25cm x 30cm) above the exhaust port of the sterilization cabinet. Portable pressure steam sterilizer uses ventilated storage box (22cm x 13cm x 6cm) instead of standard test bag. The box is filled with the test tube in the middle, the indicator is placed in the two sterilization test tubes in the middle (the test tube mouth is sealed with sterilized kraft paper), and the storage box is placed flat at the bottom of the portable pressure steam sterilizer.
After a sterilization cycle, under aseptic conditions, take out the indicator bacteria slices from the standard test bag or ventilated storage box, put them in bromocresol purple peptone water culture medium, and cultivate them at 56℃ for 7 days (self-provided biological indicator bacteria are carried out according to the instructions), and observe the color change of the culture medium. Negative control and positive control were set during detection.
(4) Judging the result: the bromocresol purple peptone water culture medium inoculated with each indicator piece does not change color, and it is judged as qualified for sterilization; When the bromocresol purple peptone water medium inoculated with one of the indicator bacteria slices changes from purple to yellow, the sterilization is unqualified.
(5) Precautions: Bacterial tablets used for monitoring must be approved by the Ministry of Health and used within the validity period.
20 .2.2 Monitoring method of dry heat sterilization effect
20.2.2. 1 chemical detection method
(1) detection method: put a chemical indicator that can indicate both the temperature and the temperature duration into the articles to be sterilized. After a sterilization cycle, take out the chemical indicator and judge whether it meets the sterilization conditions according to the changes of its color and properties.
(2) Result judgment: When the color and properties of the indicator tube are changed to the current conditions, it can be considered that the packaging has reached the sterilization conditions.
(3) Precautions: Chemical indicators used for testing shall be approved by the Ministry of Health and used within the validity period.
20.2.2.2 physical detection method: (thermocouple detection method)
(l) Detection method: During detection, the probes of multi-point temperature detectors are respectively placed at the inner, middle and outer points of each layer of the sterilizer. Close the cupboard door; Pull out the wire and observe the temperature rise and duration in the recorder.
(2) Result judgment: If the indicated temperature (curve) reaches the predetermined temperature, the sterilization temperature is qualified.
20.2.2.3 biological detection method:
(1) Indicator strain: Bacillus subtilis var. Aspergillus Niger spores (ATCC9372), the bacterial content is 5.0×105-5.0×106 cfu/cell. Its resistance should meet the following conditions: at the temperature of160 2℃, its d value is 1.3- 1.9min, the survival time is ≥3.9min, and the death time is ≤ 1.9min.
(2) Detection method: Bacillus subtilis tablets were placed in human sterile test tubes (1 tablet/tube) respectively. Two test tubes containing bacterial slices are placed in the diagonal inner corner and outer corner between the sterilizer and the door handle on each floor. The test tube cover is placed next to the test tube, and the cabinet door is closed. After a sterilization cycle, when the temperature drops to 80℃, take out the test tube after capping. Under aseptic conditions, ordinary nutrient broth medium (5ml/ tube) was added and cultured at 37℃ for 48h. The preliminary results were observed, and the sterile growth tube continued to be cultured until the seventh day.
(3) Result judgment: if the broth tube inoculated with each indicator piece is clear, it is judged that the sterilization is qualified; If the broth tube inoculated with an indicator piece is turbid, it is judged as unqualified; For broth tubes that are difficult to judge, take 0. 1ml and inoculate them on nutrient agar plates, smear them evenly with sterilized L- rods, and cultivate them at 37℃ for 48 hours. Then observe the colony morphology, do smear staining and microscopic examination to judge whether there is indicator bacteria growing. If the indicator bacteria grow, it is judged that the sterilization is unqualified; If the growth of bacteria is not indicated, it is judged that sterilization is qualified.
(4) Precautions: The indicator bacteria tablets used for testing should be approved by the Ministry of Health and used within the validity period.
20.3 Monitoring of ultraviolet disinfection effect
20.3. 1 determination of irradiance value of ultraviolet lamp
1) detection method: after turning on the ultraviolet lamp for 5 minutes, place the probe of the ultraviolet radiometer with the detection wavelength of 254nm at the center of the vertical distance 1m under the ultraviolet lamp to be detected. After the instrument is stable, the displayed data is the irradiance value of the ultraviolet lamp.
(2) Result judgment: ordinary 30w. Straight tube ultraviolet lamp, the new lamp irradiation intensity ≥90Uw/cm2 is qualified; When in use, the ultraviolet lamp irradiation intensity ≥70Uw/cm2 is qualified; The radiation intensity of the new 30W high-intensity ultraviolet lamp is ≥ 180 Uw/cm2.
(3) Precautions: When measuring, the voltage is 220v, the soil is 5v, the temperature is 20-25℃ and the relative humidity is 0.
Biological monitoring method
(1) Monitoring of air disinfection effect: Monitor according to 20.7.
① Monitoring of surface disinfection effect: Monitor according to 20.6.
20.4 Monitoring of sterilization effect of medical devices
20.4. 1 sampling time: after sterilization, samples shall be taken during the storage period.
Routine monitoring
(1) detection method: put five small medical instruments, such as suture needle, needle and surgical blade, into 5ml sterile eluent respectively; 5 pairs of syringes inhaled 5ml sterile broth for 5 times respectively; Under aseptic operation, take more than 2 large medical instruments such as surgical forceps and tweezers, scrub them repeatedly with cotton swabs stained with sterile eluent, and put the cotton swabs into 5ml of sterile eluent. Vibrate the sampling tube for 80 times, suck the lml sample to be tested with a sterile straw and put it in a sterile dish. Add15-18ml of nutrient agar melted at 45℃-48℃, pour in and shake well. After the agar was solidified, it was placed in an incubator at 37℃ for 48 hours, and the number of colonies was counted.
(2) Results: Aseptic growth on the plate met the sterilization requirements.
(3) Precautions: If the disinfection factor is a chemical disinfectant, a corresponding neutralizer should be added to the sampling solution.
20.4.3 Sterility inspection
20.4.3. 1 Preparation before sterility test
Sterile operation in 20.4.3.2: Take 5 small medical instruments, such as sewing needles, needles and blades. , and directly immersed in 6 test tubes, requires an anaerobic culture tube (one of which is a positive control) and 4 mold culture tubes. The amount of C medium is 65438 0.5 ml/tube.
Take five pairs of syringes, repeatedly suck them in 5ml eluent for five times, wash off the bacteria in the tubes, mix them evenly, and then inoculate them into anaerobic culture tubes (***6 tubes, in which 1 tube serves as a positive control) and mold culture tubes (***4 tubes). Inoculation amount: 1 ml syringe 0.5ml, 2ml syringe 1 ml, 5ml syringe 5 5- 10/0ml, 5ml syringe 20-50ml, medium amount: below 2ml,15ml/tube, 5ml, 40ml/tube.
Two large medical instruments, such as surgical forceps and tweezers, were smeared and sampled repeatedly with cotton swabs stained with sterile eluent. Put cotton swabs into sml sterile eluent, mix the sampling solution evenly, and inoculate them into anaerobic culture tube (***6 tubes, in which 1 tube is used as positive control) and mold culture medium (***4 tubes). The inoculation amount is 1 ml/tube, and the medium amount is15ml/tube.
20.4.3.3 culture: inoculate the prepared Staphylococcus aureus positive control tube solution 1: 1000 into the anaerobic culture tube of the sample to be tested, dilute 1ml, culture the anaerobic culture tube, positive control tube and negative control tube at 30-35℃ for 5 days, and culture the mold culture tube and negative control tube at 20-25℃. During the culture period, if the culture medium is turbid or precipitated after being added to the human test sample, and the appearance cannot be judged after the culture, the culture medium can be transferred to another same culture medium or slant culture medium, and after 48-72h of culture, it is observed whether there is turbidity again or whether there are colonies growing outside, and a small amount of culture medium smear is taken for dyeing, and whether there is bacterial growth is observed with a microscope.
20.4.3.4 results showed that the positive control should grow bacteria within 24 hours, and the negative control should grow aseptically during the culture period. If both anaerobic bacteria and mold culture tubes are clear or turbid, but it is proved that there is no bacterial growth, it is judged that sterilization is qualified. If any one of anaerobic bacteria and mold culture tubes 1 tube is turbid, and bacterial growth is confirmed, it should be re-sampled and re-tested twice in the same way. Except for the positive control, all other test tubes must not have bacterial growth, otherwise it will be judged as unqualified sterilization.
preventive measure
(1) The inspection time shall not exceed 6h, and if the sample is stored at 0-4℃, it shall not exceed 24h.
(2) Sample surface area
(3) If the disinfection factor is a pediatric disinfectant, a corresponding neutralizer should be added to the sampling solution.
20.4.5 pyrogen inspection method:
20.4.5. Limulus test
20.4.5.2 animal test method
Principle (1): observe the temperature rise of rabbits with a certain dose of test sample within a specified time to determine whether the pyrogen limit contained in the test sample meets the requirements.
(2) Rabbits: experimental furniture should be healthy and undamaged; Weight 1.7-3.0kg, the female rabbit should be pregnant. Seven days before the temperature is predicted, you should feed with the same feed. During this period, the weight should not be reduced, and there should be no abnormalities in spirit, appetite and excretion. Rabbits without pyrogen examination, or rabbits whose test products meet the requirements, but the temperature in the group reaches 06T; Or rabbits whose average temperature rise in the group is less than 0.8℃ and rest for more than two weeks; Or rabbits that have not been used for three weeks, their body temperature should be predicted and selected 3-7 days before checking the samples. The conditions for selecting the test are the same as those for checking the test sample; Only rabbits that don't inject liquid medicine, measure their body temperature 1 time every lh and 4 times every * * *, are within the range of 38.0-39.6℃, and the difference between the highest and lowest body temperatures is not more than 0.4℃, can be used for heat source inspection. Rabbits used for pyrogen examination should rest for at least 2 days before they can be used for the second examination if it is judged that the samples meet the requirements. If it is determined that the test sample does not meet the requirements and the average temperature rise of rabbits in the group reaches 0.8℃ or higher, all rabbits in the group will not be used, and the number of times each rabbit is used for general drug inspection shall not exceed 10.
(3) Preparation before the test: 1-2 days before the pyrogen test, the rabbits should be in the same temperature environment as far as possible, the difference between the room temperature of the laboratory and the feeding room temperature is not more than 5℃, and the laboratory temperature is 17-28℃. During the whole test, it should be noted that the change of room temperature should not be greater than 3℃, and the interference of rabbit noise should be avoided. Rabbits should stop feeding at least 1 hour before the test and put them in a suitable device until the test is completed. C an anal thermometer with an accuracy of 0.65438 0℃ or other temperature measuring devices with the same accuracy should be used to measure the temperature of rabbits. The depth and time when the anal thermometer is inserted into the human anus should be the same for each rabbit. The depth is generally about 6cm, and the time should not be less than Imin. Every 30-60 minutes, the body temperature should be measured 1 time, generally twice, and the difference between the two body temperatures should not exceed 0.2℃, and the average of the two body temperatures should be taken as the normal body temperature of the rabbit. The normal body temperature of rabbits used that day should be within the range of 38.0-39.6℃, and the difference between normal body temperatures of rabbits should not exceed l℃.
(4) Syringes, needles and all instruments in contact with the test solution should be heated in an oven at 250℃ for 30 minutes or at 65438 080℃ for 2 hours, and pyrogen can also be removed by other suitable methods.
(5) Inspection method: Take three suitable rabbits, slowly inject the specified dose of test solution from the ear vein within 65438 05 minutes after measuring their normal body temperature, and heat it to about 38℃, then measure their body temperature every lh, * * * * for three times, and subtract the normal body temperature from the highest one of the three times, which is the temperature rise degree of rabbits. If 1 rabbit's body temperature rises by 0.6℃ or above, or all three rabbits' body temperatures rise below 0.6℃, but the total increase reaches 1.4℃ or above, five rabbits should be checked again, and the inspection method is the same as above.
(6) The results showed that the temperature rise of all three rabbits in the initial test was lower than 0.6℃, and the total temperature rise of all three rabbits was lower than 65,438 0.4℃, or only 65,438 0 rabbits with temperature rise above 0.6℃ in the second test, and the total temperature rise of eight rabbits in the initial test and the second test was 3.5℃ or 3.5℃.
In the first test of three rabbits, more than 1 rabbit's body temperature increased by 0.6℃ or above, or in the second test of five rabbits; The number of rabbits whose body temperature increased by 0.6℃ or above exceeded 65438 0, or the total temperature of 8 rabbits in the initial and combined tests exceeded 3.5℃, which all considered that the pyrogen test of the test sample did not meet the requirements.
20.5 Monitoring of disinfection effect of hand and skin mucosa
20.5. 1 sampling time; Samples were taken immediately after disinfection.
sampling method
(1) Hand sampling: put the five fingers together, dip the cotton swab into the sterile eluent containing the corresponding neutralizer, rub the finger flexion surface of both hands back and forth twice (the area of one hand is about 30cm2 ~ 2), then rotate the sampling cotton swab, cut off the contact part of the operator's hand, put the cotton swab into the 10ml sterile eluent test tube containing the corresponding neutralizer, and immediately.
(2) skin mucosa sampling: the existing grid plate is sterilized with 5cm × 5crn and placed at the skin to be examined; Use 1 cotton swab without sterile eluent containing corresponding neutralizer, rub it evenly in the specification plate for 5 times, and then rotate the cotton swab. After cutting off the hand contact part, put the cotton swab into 10ml sterile eluent containing corresponding neutralizer and send it for inspection immediately. Irregular mucosal skin can be directly smeared with a cotton swab for sampling.
20.5.3 detection method
(1) Detection of the total number of bacteria: shake the sampling tube for 80 times, suck 1.0ml of the sample to be detected with a sterile pipette, inoculate it into a sterile plate, add15-18 ml of nutrient agar melted at 45-48℃, shake it evenly while pouring, and wait for the agar to solidify.
Calculation method of sampling results:
Total number of bacteria (cfu/cm2)= number of colonies on the plate × dilution multiple/sampling area (cm2)
(2) Pathogenic bacteria detection: Pathogenic bacteria detection shall be conducted according to the principle of 20 15.
20.5.4 Determination of results
(1) Disinfection and hand washing
Staff in Class L and ll areas: The total number of bacteria is ≤5Cfu/Cm2, and the disinfection is qualified if no pathogenic bacteria are detected.
Lll regional staff: The total number of bacteria is ≤ 10 Cfu/Cm2, and the disinfection is qualified if no pathogenic bacteria are detected.
Staff in lV area: the total number of bacteria is ≤ 15 Cfu/Cm2, and the disinfection is qualified if no pathogenic bacteria are detected.
No salmonella and other pathogenic bacteria were detected in the hands of the staff in the maternal and child room, the baby room, the newborn room and the pediatric ward.
(2) Skin mucosa: refer to the hygienic standard of hands.
preventive measure
In the sampling area of skin and mucosa, if the surface is less than 5cm × 5cm, the sample can be taken with the specification plate of the corresponding area.
20.6 Monitoring of disinfection effect of articles and environmental surfaces
20.6. 1 sampling time: samples after disinfection.
20.6.2 Sampling method: Place a 5cm×5cm sterilization gauge plate on the surface of the tested object, continuously sample 4 tablets, and use cotton swab 1 tablet soaked with sterile eluent containing corresponding neutralizer; Rub the cotton swab evenly back and forth in the specification plate for 5 times, and then rotate the cotton swab. After cutting off the hand contact part, put the cotton swab into a 10ml sterile eluent test tube with corresponding neutralizer, and check it immediately.
The surface of irregular objects such as door handles is directly sampled by rubbing with cotton swabs.
test method
(1) Detection of the total number of bacteria: The detection method is as per 20.5.3( 1). The result calculation of small object surface is expressed as cfu/ piece.
(2) Detection of pathogenic bacteria: The detection method is 20. 14.
Determination of results
Class L and ll area: the total number of bacteria is less than or equal to 5 cfu/cm2, and the disinfection is qualified if no pathogenic bacteria are detected.
Sick area: the total number of bacteria is ≤ 10 cfu/Cm2, and the disinfection is qualified if no pathogenic bacteria are detected.
LV area: the total number of bacteria is ≤ 15 cfu/Cm2, no pathogenic bacteria are found, and the disinfection is qualified.
Salmonella shall not be detected on the surface of articles in maternal and infant room, premature baby room, baby room, newborn room and pediatric ward.
preventive measure
(1) delivery time: according to 20.4.4( 1).
(2) sampling area; Press 20.4.4(2) for execution.
20.7 Monitoring of air disinfection effect
20.7. 1 sampling time: sampling after disinfection and before operation.
sampling method
(1) layout method; The indoor area is ≤ 30cm ~ 2, and there are three diagonal points on the inner, middle and outer surfaces, and the distribution points of the inner and outer points are away from the wall 1m; Indoor area > 30㎡, 4 corners and 5 points in the center, and the distribution position of 4 corners is away from the wall1m.
(2) Plate exposure method: Place ordinary nutrient agar plates (9CM in diameter) at indoor sampling points, and the sampling height is 1.5m from the ground. When sampling, open the plate cover, buckle it next to the plate, expose it for 5 minutes, and immediately cover it for inspection.
20.7.3 Detection method: Place the plate to be detected in an incubator at 37℃ for 48h, count the number of colonies and isolate pathogenic bacteria.