CN105675604A - Method for detecting antibiotic residues in milk - Google Patents
Method for detecting antibiotic residues in milk Download PDFInfo
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- CN105675604A CN105675604A CN201610194687.6A CN201610194687A CN105675604A CN 105675604 A CN105675604 A CN 105675604A CN 201610194687 A CN201610194687 A CN 201610194687A CN 105675604 A CN105675604 A CN 105675604A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N21/78—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour
Abstract
The invention discloses a method for detecting antibiotic residues in milk. The method comprises the specific steps of inoculating strains of bacillus subtilis on an inclined culture medium for optimized cultivation; performing successive transfer culture and collection; washing bacterial lawns with normal saline, transferring the bacterial lawns into a germ-free container, fully oscillating the bacterial lawns, and filtering to prepare suspension prepared into bacterial sludge; adding an agar indicator solution, and placing reaction liquid into each hole of a pore plate or a trace reactor; adding a milk sample into each hole of a kit or the trace reactor, carefully sealing the kit or the trace reactor with an aluminum-plastic film, and checking whether the liquid discolors after constant-temperature reaction is generated in a water bath or a constant-temperature box. According to the method, the strains of the bacillus subtilis are stable and easy to store and prepare and can bear heat in a preparation process; even under an extensive storage condition, the strains are difficult to inactivate; the detection method is easy and convenient to operate, high in sensitivity and low in cost; quick detection and high-flux detection are realized without expensive detection equipment, and the detection limit is low.
Description
Technical field
The present invention relates to antibiotics leftover detection technical field, specifically the detection method of a kind of Residue of Antibiotics in Milk.
Background technology
All nutritive substances that the needed by human body that milk is rich in except food fibre is wanted, the height of its nutrition is that other foods are irreplaceable. Fresh milk and the own important component part through becoming in people (especially old man and children) life food of milk preparation. In recent years, along with developing rapidly of China's livestock breeding industry, milk yield increases substantially, in order to generation and the treatment disease such as mastitis of preventing disease, cultivation person can add the fodder additives containing certain proportion microbiotic in ox feed and in tap water, but, will unavoidably cause the residual of microbiotic in milk like this. If long-term drinking contains the milk of microbiotic, the microbiotic in milk enters human body by food chain, equals beyond doubt to take low dose of microbiotic for a long time. Anaphylaxis can be caused after the people of microbiotic allergic constitution is taken the milk of residual antibiotic. Normally drink user, the conditionality pathogenic bacterium of some in body can be caused easily to produce resistance, be difficult to prove effective with antibiotic therapy of the same race again once ill. Not only harm humans is healthy for antibiotic remains, also affects the quality of milk simultaneously. The most high residue amount of microbiotic in milk has been made detailed regulation by the food law of the World Food Programme, the World Health Organization. The Ministry of Agriculture of China has issued and implemented " the fresh cow's milk industry standard of pollution-free food " for 2001. Therefore, set up and a kind of detect the method for multiple antibiotic residues in milk simply, quickly and efficiently and be very important.
Due to the medicine of sulfamido, quinolones, beta-lactam, tetracyclines, chloromycetin totally 5 big class 33 kinds of veterinary drugs, its chemical molecular structure and chemical property differ greatly, detection while multiple antibiotic residues in milk there is is certain difficulty simultaneously, also do not have document to report at present. For Residue of Antibiotics in Milk analysis, the pretreatment technologies such as solid phase extraction techniques, solid phase micro-extraction technique and matrix solid phase dispersion have been applied in sample pre-treatments step, to be used for the matrix interference problem solved in sample analysis test. Wherein, being most widely used of solid phase extraction techniques, but be applied in the pre-treatment of Residue of Antibiotics in Milk detection, solid phase extraction techniques but generally exist sample handling processes complexity, toxic reagent many, the consumptive material consumption of contact greatly, need that extra nitrogen blows, poor reproducibility and the automatically problem such as online cannot be realized. The particularly medicine such as tetracyclines, owing to it is combined with protein or mineral substance, forms combined, has therefore more increased the weight of the detection difficulty of residual antibiotic in milk.
Summary of the invention
It is an object of the invention to provide the detection method that a kind of cost is low, detect fireballing Residue of Antibiotics in Milk, to solve in above-mentioned background technology the problem proposed.
For achieving the above object, the present invention provides following technical scheme:
A detection method for Residue of Antibiotics in Milk, concrete steps are as follows:
(1) by the inoculation of subtilis on slant medium, the pH value of adjustable inclined surface apparatus substratum is 6.5-7, being optimized at 45-65 DEG C and cultivate 48-72h, described slant medium comprises according to the raw material of weight part: peptone 2-4 part, extractum carnis powder 2.5-3.5 part, yeast extract paste 5-6 part, agar 10-20 part, sodium-chlor 0.4-0.6 part, glucose 1-2 part, dipotassium hydrogen phosphate 0.01-0.03 part, distilled water 100-120 part;
(2) it is 7-8 by the pH value of sodium hydroxide solution adjustable inclined surface apparatus substratum, collect after carrying out transferred species succeeding transfer culture 24-36h, lower bacterium tongue is washed again with physiological saline, it is transferred in sterile chamber, fully after vibration, filter, make suspension, hot suspension 8-12min is added at 60-70 DEG C, after hold over night, with the centrifugation 15-25min of 1500-1700r/min, get supernatant liquor, the centrifugation 5-10min of 3800-4000r/min again, abandoning supernatant, after centrifuge washing repeatedly like this 3-5 time, with physiological saline, centrifugation suspended into subtilis bacterium mud, adjust its OD600 value to 0.25-0.35, store at 0-4 DEG C, for subsequent use,
(3) it is 3-4: 1 add agar indicator solution by the volume ratio of agar indicator solution and subtilis bacterium mud, mix at 60-70 DEG C, obtain reaction solution, in each hole be respectively charged into orifice plate by described reaction solution 0.2-0.4mL or microscale reactor, the subtilis in every hole or microscale reactor reaches 0.5-1.5 × 108Cfu, covers aluminum plastic film and carries out heat sealing, after being inverted 20-40min in 60-70 DEG C, is cooled to normal temperature along putting, and loads aluminium plastic bag, and protection against the tide, with keeping in Dark Place, makes detection kit; Described agar indicator solution comprises according to the raw material of weight part: agar 1.5-3.5 part, purpurum bromocresolis 0.0008-0.0012 part, sodium hydroxide 0.02-0.04 part, peptone 1.0-1.5 part, extractum carnis powder 2.5-3.5 part, sodium-chlor 0.4-0.6 part, distilled water 93-95 part;
(4) in every hole or microscale reactor of the described test kit obtained, milk sample 0.05-0.25mL is added, carefully seal with aluminum plastic film, isothermal reaction 2.3-2.5h in the water-bath of 50-60 DEG C or thermostat container, the every hole of the test kit of negative sample or microscale reactor become yellow by blueness, color change interval pH is 5.0-6.0, represents that antibiotic remains is negative; Positive nondiscoloration, represents that antibiotic remains is positive.
As the scheme that the present invention is further: described slant medium comprises according to the raw material of weight part: peptone 2.5-3.5 part, extractum carnis powder 2.8-3.2 part, yeast extract paste 5.2-5.8 part, agar 12-18 part, sodium-chlor 0.45-0.55 part, glucose 1.2-1.8 part, dipotassium hydrogen phosphate 0.015-0.025 part, distilled water 105-115 part.
As the scheme that the present invention is further: described slant medium comprises according to the raw material of weight part: peptone 3 parts, 3 parts, extractum carnis powder, yeast extract paste 5.5 parts, 15 parts, agar, 0.5 part, sodium-chlor, glucose 1.5 parts, dipotassium hydrogen phosphate 0.02 part, distilled water 110 parts.
As the scheme that the present invention is further: described agar indicator solution comprises according to the raw material of weight part: agar 2-3 part, purpurum bromocresolis 0.0009-0.0011 part, sodium hydroxide 0.025-0.035 part, peptone 1.1-1.4 part, extractum carnis powder 2.8-3.2 part, sodium-chlor 0.45-0.55 part, distilled water 93.5-94.5 part.
As the scheme that the present invention is further: described agar indicator solution comprises according to the raw material of weight part: 2.5 parts, agar, purpurum bromocresolis 0.001 part, 0.03 part, sodium hydroxide, peptone 1.2 parts, 3 parts, extractum carnis powder, 0.5 part, sodium-chlor, distilled water 94 parts.
As the present invention's further scheme: in described step (2), the concentration of sodium hydroxide solution is 1-2mol/L.
Compared with prior art, the invention has the beneficial effects as follows:
In the present invention, the bacterial strain of subtilis is stablized, and easily preserves and preparation, can tolerate the heat in the course of processing, also not easy in inactivation under extensive preservation condition, easy and simple to handle, highly sensitive, cost is low, detect fast, high throughput testing, it is not necessary to expensive test set, detectability is low.
Embodiment
Below in conjunction with embodiment, the technical scheme of this patent is described in more detail.
Embodiment 1
A detection method for Residue of Antibiotics in Milk, concrete steps are as follows:
(1) by the inoculation of subtilis on slant medium, the pH value of adjustable inclined surface apparatus substratum is 6.5, being optimized at 45 DEG C and cultivate 48h, described slant medium comprises according to the raw material of weight part: peptone 2 parts, 2.5 parts, extractum carnis powder, yeast extract paste 5 parts, 10 parts, agar, 0.4 part, sodium-chlor, glucose 1 part, dipotassium hydrogen phosphate 0.01 part, distilled water 100 parts;
(2) employing concentration is the pH value of the sodium hydroxide solution adjustable inclined surface apparatus substratum of 1mol/L is 7, collect after carrying out transferred species succeeding transfer culture 24h, lower bacterium tongue is washed again with physiological saline, it is transferred in sterile chamber, fully after vibration, filter, make suspension, hot suspension 8min is added at 60 DEG C, after hold over night, with the centrifugation 15min of 1500r/min, get supernatant liquor, the centrifugation 5min of 3800r/min again, abandoning supernatant, after centrifuge washing repeatedly like this 3 times, with physiological saline, centrifugation suspended into subtilis bacterium mud, adjust its OD600 value to 0.25, store at 0 DEG C, for subsequent use,
(3) it is add agar indicator solution at 3: 1 by the volume ratio of agar indicator solution and subtilis bacterium mud, mix at 60 DEG C, obtain reaction solution, in each hole be respectively charged into orifice plate by described reaction solution 0.2mL or microscale reactor, the subtilis in every hole or microscale reactor reaches 0.5 × 108Cfu, covers aluminum plastic film and carries out heat sealing, after being inverted 20min in 60 DEG C, is cooled to normal temperature along putting, and loads aluminium plastic bag, and protection against the tide, with keeping in Dark Place, makes detection kit; Described agar indicator solution comprises according to the raw material of weight part: 1.5 parts, agar, purpurum bromocresolis 0.0008 part, 0.02 part, sodium hydroxide, peptone 1.0 parts, 2.5 parts, extractum carnis powder, 0.4 part, sodium-chlor, distilled water 93 parts;
(4) in every hole or microscale reactor of the described test kit obtained, milk sample 0.05mL is added, carefully seal with aluminum plastic film, isothermal reaction 2.3h in the water-baths of 50 DEG C or thermostat container, the every hole of the test kit of negative sample or microscale reactor become yellow by blueness, color change interval pH is 5.0, represents that antibiotic remains is negative; Positive nondiscoloration, represents that antibiotic remains is positive.
Embodiment 2
A detection method for Residue of Antibiotics in Milk, concrete steps are as follows:
(1) by the inoculation of subtilis on slant medium, the pH value of adjustable inclined surface apparatus substratum is 6.6, being optimized at 50 DEG C and cultivate 56h, described slant medium comprises according to the raw material of weight part: peptone 2.8 parts, 2.9 parts, extractum carnis powder, yeast extract paste 5.3 parts, 13 parts, agar, 0.48 part, sodium-chlor, glucose 1.3 parts, dipotassium hydrogen phosphate 0.018 part, distilled water 108 parts;
(2) employing concentration is the pH value of the sodium hydroxide solution adjustable inclined surface apparatus substratum of 1.2mol/L is 7.2, collect after carrying out transferred species succeeding transfer culture 28h, lower bacterium tongue is washed again with physiological saline, it is transferred in sterile chamber, fully after vibration, filter, make suspension, hot suspension 9min is added at 62 DEG C, after hold over night, with the centrifugation 18min of 1550r/min, get supernatant liquor, the centrifugation 6min of 3850r/min again, abandoning supernatant, after centrifuge washing repeatedly like this 4 times, with physiological saline, centrifugation suspended into subtilis bacterium mud, adjust its OD600 value to 0.28, store at 1 DEG C, for subsequent use,
(3) it is add agar indicator solution at 3.2: 1 by the volume ratio of agar indicator solution and subtilis bacterium mud, mix at 62 DEG C, obtain reaction solution, in each hole be respectively charged into orifice plate by described reaction solution 0.25mL or microscale reactor, the subtilis in every hole or microscale reactor reaches 0.8 × 108Cfu, covers aluminum plastic film and carries out heat sealing, after being inverted 25min in 62 DEG C, is cooled to normal temperature along putting, and loads aluminium plastic bag, and protection against the tide, with keeping in Dark Place, makes detection kit; Described agar indicator solution comprises according to the raw material of weight part: 2.2 parts, agar, purpurum bromocresolis 0.00095 part, 0.028 part, sodium hydroxide, peptone 1.15 parts, 2.9 parts, extractum carnis powder, 0.48 part, sodium-chlor, distilled water 93.8 parts;
(4) in every hole or microscale reactor of the described test kit obtained, milk sample 0.1mL is added, carefully seal with aluminum plastic film, isothermal reaction 2.35h in the water-baths of 52 DEG C or thermostat container, the every hole of the test kit of negative sample or microscale reactor become yellow by blueness, color change interval pH is 5.2, represents that antibiotic remains is negative; Positive nondiscoloration, represents that antibiotic remains is positive.
Embodiment 3
A detection method for Residue of Antibiotics in Milk, concrete steps are as follows:
(1) by the inoculation of subtilis on slant medium, the pH value of adjustable inclined surface apparatus substratum is 6.8, being optimized at 55 DEG C and cultivate 51h, described slant medium comprises according to the raw material of weight part: peptone 3 parts, 3 parts, extractum carnis powder, yeast extract paste 5.5 parts, 15 parts, agar, 0.5 part, sodium-chlor, glucose 1.5 parts, dipotassium hydrogen phosphate 0.02 part, distilled water 110 parts;
(2) employing concentration is the pH value of the sodium hydroxide solution adjustable inclined surface apparatus substratum of 1.5mol/L is 7.5, collect after carrying out transferred species succeeding transfer culture 31h, lower bacterium tongue is washed again with physiological saline, it is transferred in sterile chamber, fully after vibration, filter, make suspension, hot suspension 10min is added at 65 DEG C, after hold over night, with the centrifugation 20min of 1600r/min, get supernatant liquor, the centrifugation 8min of 3900r/min again, abandoning supernatant, after centrifuge washing repeatedly like this 5 times, with physiological saline, centrifugation suspended into subtilis bacterium mud, adjust its OD600 value to 0.3, store at 2 DEG C, for subsequent use,
(3) it is add agar indicator solution at 3.5: 1 by the volume ratio of agar indicator solution and subtilis bacterium mud, mix at 65 DEG C, obtain reaction solution, in each hole be respectively charged into orifice plate by described reaction solution 0.3mL or microscale reactor, the subtilis in every hole or microscale reactor reaches 1 × 108Cfu, covers aluminum plastic film and carries out heat sealing, after being inverted 30min in 65 DEG C, is cooled to normal temperature along putting, and loads aluminium plastic bag, and protection against the tide, with keeping in Dark Place, makes detection kit;Described agar indicator solution comprises according to the raw material of weight part: 2.5 parts, agar, purpurum bromocresolis 0.001 part, 0.03 part, sodium hydroxide, peptone 1.2 parts, 3 parts, extractum carnis powder, 0.5 part, sodium-chlor, distilled water 94 parts;
(4) in every hole or microscale reactor of the described test kit obtained, milk sample 0.15mL is added, carefully seal with aluminum plastic film, isothermal reaction 2.4h in the water-baths of 55 DEG C or thermostat container, the every hole of the test kit of negative sample or microscale reactor become yellow by blueness, color change interval pH is 5.5, represents that antibiotic remains is negative; Positive nondiscoloration, represents that antibiotic remains is positive.
Embodiment 4
A detection method for Residue of Antibiotics in Milk, concrete steps are as follows:
(1) by the inoculation of subtilis on slant medium, the pH value of adjustable inclined surface apparatus substratum is 6.9, being optimized at 60 DEG C and cultivate 56h, described slant medium comprises according to the raw material of weight part: peptone 3.5 parts, 3.2 parts, extractum carnis powder, yeast extract paste 5.8 parts, 18 parts, agar, 0.55 part, sodium-chlor, glucose 1.8 parts, dipotassium hydrogen phosphate 0.025 part, distilled water 115 parts;
(2) employing concentration is the pH value of the sodium hydroxide solution adjustable inclined surface apparatus substratum of 1.8mol/L is 7.8, collect after carrying out transferred species succeeding transfer culture 34h, lower bacterium tongue is washed again with physiological saline, it is transferred in sterile chamber, fully after vibration, filter, make suspension, hot suspension 11min is added at 68 DEG C, after hold over night, with the centrifugation 22min of 1650r/min, get supernatant liquor, the centrifugation 9min of 3950r/min again, abandoning supernatant, after centrifuge washing repeatedly like this 3 times, with physiological saline, centrifugation suspended into subtilis bacterium mud, adjust its OD600 value to 0.32, store at 3 DEG C, for subsequent use,
(3) it is add agar indicator solution at 3.8: 1 by the volume ratio of agar indicator solution and subtilis bacterium mud, mix at 68 DEG C, obtain reaction solution, in each hole be respectively charged into orifice plate by described reaction solution 0.35mL or microscale reactor, the subtilis in every hole or microscale reactor reaches 1.2 × 108Cfu, covers aluminum plastic film and carries out heat sealing, after being inverted 35min in 68 DEG C, is cooled to normal temperature along putting, and loads aluminium plastic bag, and protection against the tide, with keeping in Dark Place, makes detection kit; Described agar indicator solution comprises according to the raw material of weight part: 3 parts, agar, purpurum bromocresolis 0.0011 part, 0.035 part, sodium hydroxide, peptone 1.4 parts, 3.2 parts, extractum carnis powder, 0.55 part, sodium-chlor, distilled water 94.5 parts;
(4) in every hole or microscale reactor of the described test kit obtained, milk sample 0.2mL is added, carefully seal with aluminum plastic film, isothermal reaction 2.45h in the water-baths of 58 DEG C or thermostat container, the every hole of the test kit of negative sample or microscale reactor become yellow by blueness, color change interval pH is 5.8, represents that antibiotic remains is negative; Positive nondiscoloration, represents that antibiotic remains is positive.
Embodiment 5
A detection method for Residue of Antibiotics in Milk, concrete steps are as follows:
(1) by the inoculation of subtilis on slant medium, the pH value of adjustable inclined surface apparatus substratum is 7, being optimized at 65 DEG C and cultivate 72h, described slant medium comprises according to the raw material of weight part: peptone 4 parts, 3.5 parts, extractum carnis powder, yeast extract paste 6 parts, 20 parts, agar, 0.6 part, sodium-chlor, glucose 2 parts, dipotassium hydrogen phosphate 0.03 part, distilled water 120 parts;
(2) employing concentration is the pH value of the sodium hydroxide solution adjustable inclined surface apparatus substratum of 2mol/L is 8, collect after carrying out transferred species succeeding transfer culture 36h, lower bacterium tongue is washed again with physiological saline, it is transferred in sterile chamber, fully after vibration, filter, make suspension, hot suspension 12min is added at 70 DEG C, after hold over night, with the centrifugation 25min of 1700r/min, get supernatant liquor, the centrifugation 10min of 4000r/min again, abandoning supernatant, after centrifuge washing repeatedly like this 5 times, with physiological saline, centrifugation suspended into subtilis bacterium mud, adjust its OD600 value to 0.35, store at 4 DEG C, for subsequent use,
(3) it is add agar indicator solution at 4: 1 by the volume ratio of agar indicator solution and subtilis bacterium mud, mix at 70 DEG C, obtain reaction solution, in each hole be respectively charged into orifice plate by described reaction solution 0.4mL or microscale reactor, the subtilis in every hole or microscale reactor reaches 1.5 × 108Cfu, covers aluminum plastic film and carries out heat sealing, after being inverted 40min in 70 DEG C, is cooled to normal temperature along putting, and loads aluminium plastic bag, and protection against the tide, with keeping in Dark Place, makes detection kit; Described agar indicator solution comprises according to the raw material of weight part: 3.5 parts, agar, purpurum bromocresolis 0.0012 part, 0.04 part, sodium hydroxide, peptone 1.5 parts, 3.5 parts, extractum carnis powder, 0.6 part, sodium-chlor, distilled water 95 parts;
(4) in every hole or microscale reactor of the described test kit obtained, milk sample 0.25mL is added, carefully seal with aluminum plastic film, isothermal reaction 2.5h in the water-baths of 60 DEG C or thermostat container, the every hole of the test kit of negative sample or microscale reactor become yellow by blueness, color change interval pH is 6.0, represents that antibiotic remains is negative; Positive nondiscoloration, represents that antibiotic remains is positive.
In the present invention, the bacterial strain of subtilis is stablized, and easily preserves and preparation, can tolerate the heat in the course of processing, also not easy in inactivation under extensive preservation condition, easy and simple to handle, highly sensitive, cost is low, detect fast, high throughput testing, it is not necessary to expensive test set, detectability is low.
Above the better embodiment of this patent is explained in detail, but this patent is not limited to above-mentioned enforcement mode, in the ken that one skilled in the relevant art possesses, it is also possible under the prerequisite not departing from this patent objective, make various change.
Claims (6)
1. the detection method of a Residue of Antibiotics in Milk, it is characterised in that, concrete steps are as follows:
(1) by the inoculation of subtilis on slant medium, the pH value of adjustable inclined surface apparatus substratum is 6.5-7, being optimized at 45-65 DEG C and cultivate 48-72h, described slant medium comprises according to the raw material of weight part: peptone 2-4 part, extractum carnis powder 2.5-3.5 part, yeast extract paste 5-6 part, agar 10-20 part, sodium-chlor 0.4-0.6 part, glucose 1-2 part, dipotassium hydrogen phosphate 0.01-0.03 part, distilled water 100-120 part;
(2) it is 7-8 by the pH value of sodium hydroxide solution adjustable inclined surface apparatus substratum, collect after carrying out transferred species succeeding transfer culture 24-36h, lower bacterium tongue is washed again with physiological saline, it is transferred in sterile chamber, fully after vibration, filter, make suspension, hot suspension 8-12min is added at 60-70 DEG C, after hold over night, with the centrifugation 15-25min of 1500-1700r/min, get supernatant liquor, the centrifugation 5-10min of 3800-4000r/min again, abandoning supernatant, after centrifuge washing repeatedly like this 3-5 time, with physiological saline, centrifugation suspended into subtilis bacterium mud, adjust its OD600 value to 0.25-0.35, store at 0-4 DEG C, for subsequent use,
(3) it is 3-4: 1 add agar indicator solution by the volume ratio of agar indicator solution and subtilis bacterium mud, mix at 60-70 DEG C, obtain reaction solution, in each hole be respectively charged into orifice plate by described reaction solution 0.2-0.4mL or microscale reactor, the subtilis in every hole or microscale reactor reaches 0.5-1.5 × 108Cfu, covers aluminum plastic film and carries out heat sealing, after being inverted 20-40min in 60-70 DEG C, is cooled to normal temperature along putting, and loads aluminium plastic bag, and protection against the tide, with keeping in Dark Place, makes detection kit; Described agar indicator solution comprises according to the raw material of weight part: agar 1.5-3.5 part, purpurum bromocresolis 0.0008-0.0012 part, sodium hydroxide 0.02-0.04 part, peptone 1.0-1.5 part, extractum carnis powder 2.5-3.5 part, sodium-chlor 0.4-0.6 part, distilled water 93-95 part;
(4) in every hole or microscale reactor of the described test kit obtained, milk sample 0.05-0.25mL is added, carefully seal with aluminum plastic film, isothermal reaction 2.3-2.5h in the water-bath of 50-60 DEG C or thermostat container, the every hole of the test kit of negative sample or microscale reactor become yellow by blueness, color change interval pH is 5.0-6.0, represents that antibiotic remains is negative; Positive nondiscoloration, represents that antibiotic remains is positive.
2. the detection method of Residue of Antibiotics in Milk according to claim 1, it is characterized in that, described slant medium comprises according to the raw material of weight part: peptone 2.5-3.5 part, extractum carnis powder 2.8-3.2 part, yeast extract paste 5.2-5.8 part, agar 12-18 part, sodium-chlor 0.45-0.55 part, glucose 1.2-1.8 part, dipotassium hydrogen phosphate 0.015-0.025 part, distilled water 105-115 part.
3. the detection method of Residue of Antibiotics in Milk according to claim 1, it is characterized in that, described slant medium comprises according to the raw material of weight part: peptone 3 parts, 3 parts, extractum carnis powder, yeast extract paste 5.5 parts, 15 parts, agar, 0.5 part, sodium-chlor, glucose 1.5 parts, dipotassium hydrogen phosphate 0.02 part, distilled water 110 parts.
4. the detection method of Residue of Antibiotics in Milk according to claim 1, it is characterized in that, described agar indicator solution comprises according to the raw material of weight part: agar 2-3 part, purpurum bromocresolis 0.0009-0.0011 part, sodium hydroxide 0.025-0.035 part, peptone 1.1-1.4 part, extractum carnis powder 2.8-3.2 part, sodium-chlor 0.45-0.55 part, distilled water 93.5-94.5 part.
5. the detection method of Residue of Antibiotics in Milk according to claim 1, it is characterized in that, state agar indicator solution and comprise according to the raw material of weight part: 2.5 parts, agar, purpurum bromocresolis 0.001 part, 0.03 part, sodium hydroxide, peptone 1.2 parts, 3 parts, extractum carnis powder, 0.5 part, sodium-chlor, distilled water 94 parts.
6. the detection method of Residue of Antibiotics in Milk according to claim 1, it is characterised in that, in described step (2), the concentration of sodium hydroxide solution is 1-2mol/L.
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| CN106319026A (en) * | 2016-08-19 | 2017-01-11 | 界首市科创食品检测中心 | Method for detecting antibiotic in milk |
| CN106867940A (en) * | 2017-03-16 | 2017-06-20 | 青岛农业大学 | A kind of method for promoting the growth of bacillus stearothermophilus gemma to sprout |
| CN107314980A (en) * | 2017-06-08 | 2017-11-03 | 中国肉类食品综合研究中心 | Multiple antibiotic residues detection kit and detection method in a kind of animal derived food |
| CN107917911A (en) * | 2017-11-15 | 2018-04-17 | 中华人民共和国龙口出入境检验检疫局 | The remaining method of antibiotic in one kind detection meat product |
| CN108414639A (en) * | 2018-03-27 | 2018-08-17 | 山东五洲检测有限公司 | A kind of rapid detection method of Pesticide Residues object |
| CN109270056A (en) * | 2018-10-12 | 2019-01-25 | 中国肉类食品综合研究中心 | A kind of method and microorganism colour reagent box detecting high-throughput antibiotic residue |
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| CN106867940B (en) * | 2017-03-16 | 2020-07-07 | 青岛农业大学 | Method for promoting growth and germination of bacillus stearothermophilus spores |
| CN107314980A (en) * | 2017-06-08 | 2017-11-03 | 中国肉类食品综合研究中心 | Multiple antibiotic residues detection kit and detection method in a kind of animal derived food |
| CN107314980B (en) * | 2017-06-08 | 2020-02-18 | 中国肉类食品综合研究中心 | Detection kit and detection method for multiple antibiotic residues in animal-derived food |
| CN107917911A (en) * | 2017-11-15 | 2018-04-17 | 中华人民共和国龙口出入境检验检疫局 | The remaining method of antibiotic in one kind detection meat product |
| CN108414639A (en) * | 2018-03-27 | 2018-08-17 | 山东五洲检测有限公司 | A kind of rapid detection method of Pesticide Residues object |
| CN109270056A (en) * | 2018-10-12 | 2019-01-25 | 中国肉类食品综合研究中心 | A kind of method and microorganism colour reagent box detecting high-throughput antibiotic residue |
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