LSY-10059 Spectinomycin ELISA Test Kit for milk, honey, tissue

Spectinomycin ELISA Test Kit

Catalog No. LSY-10059

1. Principle

This test kit is based on the competitive enzyme immunoassay for the detection of Spectinmycin in the sample. The coupling antigens are pre-coated on the micro-well stripes. The Spectinmycin in the sample and the coupling antigens pre-coated on the micro-well stripes compete for the anti-Spectinomycin antibodies. After the addition of the enzyme conjugate, the TMB substrate is added for coloration. The optical density (OD) value of the sample has a negative correlation with the Spectinomycin in it. This value is compared to the standard curve and the Spectinomycin concentration is subsequently obtained.

2. Technical specifications

Sensitivity: 0.3ppb

Incubation Temperature: 25℃

Incubation Time: 20min～10min

Detection limit

Liquid milk about 5ppb

Tissue about 2ppb

Honey about 1ppb

Note: ppb= ng/ml or ng/g

Cross-reaction rate

Spectinomycin 100%

Tilmicosin <0.1%

Lincomycin <0.1%
Kanamycin <0.1%

Gentamicin <0.1%
Apramycin <0.1%
Neomycin <0.1%

Streptomycin <0.1%

Recovery rate

Tissue 90±30%

3. Components

• Micro-well strips: 12 strips with 8 removable wells each
• 6× standard solution (1mL each): 0ppb, 0.3ppb, 0.9ppb, 2.7ppb,8.1 ppb
• Enzyme conjugate (6 mL)
• Antibody working solution (6 mL)
• Substrate A (7 mL)
• Substrate B (7 mL)
• Stop solution (6 mL)
• 20X concentrated washing buffer (30 mL)
• 10X concentrated redissolving solution (20 mL)

4. Materials required but not provided

• Equipments: microplate reader, printer, homogenizer, nitrogen-drying device, vortex, centrifuge, measuring pipets, balance (a reciprocal sensibility of 0.01 g), incubator, water bath;
• Micropipettors: single-channel 20-200µL, 100-1000µL, and multi-channel 30～300µl;
• Reagents: Deionized water, HCl

5. Sample pre-treatment

Instructions

The following points must be dealt with before the pre-treatment of any kind of sample:

• Only the disposable tips can be used for the experiments and the tips must be changed when used for absorbing different reagents;
• Before the experiment, each experimental equipment must be clean and should be re-cleaned if necessary, in order to avoid the contamination that interferes with the experimental results.

Solution preparation before sample pre-treatment:

• Redissolving solution: dissolve 1 part of 10x concentrated redissoving solution in 9 part of deionized water at the ratio of 1: 9.
• Washing buffer: dissolve 1 part of 20x concentrated washing buffer with 19 part of deionized water at the ratio of 1:19.
• 1M HCl: take 1ml HCl, add 11ml deionized water, mix it evenly.

5.1 Samples preparation

a) Milk

1) Take the collected raw milk , thaw and return to room temperature for above 30min;

2) Put tips into down-layer of raw milk, take out 1ml sample into 2ml centrifuge tube(Note: do not take up-layer cream);

3) Add 50ul 1M HCl, shake strongly for 1min (or vortex for 30s);

4) Centrifuge at above 4000 g at room temperature for 10min;

5) Take 50ul up-layer clear liquid into another centrifuge tube (do not take up-layer cream), then add 450ul Tissue Redissolving solution,shake strongly for 1min (or vortex for 30s);

6) Take 50ul liquid to test.

Fold of dilution of the sample: 10

b) Tissue (chicken, pork)

1) Weigh 1± 0.05g of the homogenized tissue sample, add into 50mL of the plastic centrifuge tube.

2) Add 2mL Redissolving solution, shake strongly for 2min(or vortex for 1min), then Centrifuge at above 4000r/min at room temperature (20-25 ℃) for 10min.

• Take 50 µL of the up-layer clean liquid for analysis.

Fold of dilution of the sample:2

c) Honey

1) Weigh 1± 0.05g honey sample, add into 50mL of the plastic centrifuge tube;

2) Add 2mL deionized water, shake strongly for 2 min(or vortex for 2min), or use centrifuge for 2 min.

• Add 100µL supernatant, add 400ul Redissolving solution, vortex for 30s.
• Take 50 µL for analysis.

Fold of dilution of the sample: 10