LSY-10067 Cefalexin ELISA Test Kit for milk, egg, tissue sample

Cefalexin ELISA Test Kit

Catalog No. LSY-10067

1. Principle

This test kit is based on the indirect competitive enzyme immunoassay for the detection of Cefalexin in samples. The coupling antigens are pre-coated on the micro-well stripes. The Cefalexin in the sample and the coupling antigens pre-coated on the micro-well stripes compete for the anti-Cefalexin 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 Cefalexin in the sample. This value is compared to the standard curve and the Cefalexin concentration is subsequently obtained.

2. Technical specifications

Sensitivity: 0.3 ppb

Incubation Temperature: 25℃

Incubation Time: 30min—15min

Detection limit:

Tissue 10ppb

Egg 15ppb

Liquid milk 5ppb

Milk powder 24ppb

Cross-reaction rate:

Cefalexin 100%

Recovery rate

Tissue 100%±30%

Egg, liquid milk, milk powder 90%±30%

3. Components

4. Materials required but not provided

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

5. Sample pre-treatment

Instructions (The following points must be dealt with before the pre-treatment)

1) 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 utensil 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:

• 1M NaOH: Take 4.0g NaOH, add deionized water to 100mL, mix it evenly.
• 10% Methanol: 1 part of Methanol + 9 parts of deionized water, mix it evenly.
• The 10× concentrated redissolving solution is diluted with deionized water at 1:9 (1 part of 10x concentrated redissolving solution + 9 parts of deionized water), mix it evenly.
• Extracting buffer A: 1 part of 20× extracting buffer A + 19 parts of deionized water, mix it evenly.
• Extracting buffer B: 1 part of 2× extracting buffer B + 1 part of deionized water, mix it evenly.

5.1 Tissue

1) Weigh 2.0 ± 0.05 g of the homogenized tissue sample into 50 ml centrifuge tube, add 3ml Extracting buffer A, vortex for 3min;

2) Then add 600µl 1M NaOH and 2.4ml Extracting buffer B, vortex for 3 min, centrifuge at above 4000 r/min at room temperature for 5 min;

3) Take 100µl up-layer liquid, add 400µl diluted redissolving solution, mix it evenly;

4) Take 50µl solution for further analysis.

Fold of dilution of the sample: 20

5.2 Egg

1) Weigh 1±0.05g egg sample into 10ml centrifuge tube, add 5ml 10% Methanol, vortex for 3min, centrifuge at above 4000 r/min at room temperature for 5 min; take 100ul up-layer liquid, add 900µl diluted redissolving solution, shake for 30s;

2) Take 50µl solution for further analysis.

Fold of dilution of the sample: 50

5.3 Liquid milk

• Take 100µl liquid milk into 2ml centrifuge tube, add 900µl diluted redissolving solution, vortex for 30s, centrifuge at above 4000 r/min at room temperature for 5 min;
• Take 50 µL up-layer liquid for further analysis

Fold of dilution of the sample: 10

5.4 Milk powder

1) Weigh 1.0 ± 0.05 g of the milk powder sample into 10 ml centrifuge tube, add 7ml deionized water, vortex for 3min, centrifuge at above 4000 r/min at room temperature for 5 min;

2) Take 100µl up-layer liquid into 2ml centrifuge tube, add 900µl diluted redissolving solution, vortex for 30s,centrifuge at above 4000 r/min at room temperature for 5 min; ;

4) Take 50µl up-layer liquid for further analysis.

Fold of dilution of the sample: 80