Human Apolipoprotein B
ELISA development kit
Product Code: 3715-1A-20

CONTENTS, development kit for 20 plates:
Vial 1 (blue top)
Monoclonal antibody LDL 20/17 (500 μl)
Concentration: 1 mg/ml
Vial 2 (red top)
Biotinylated monoclonal antibody LDL 11 (250 μl)
Concentration: 1 mg/ml
Vial 3 (white top)
Streptavidin-Alkaline Phosphatase (250 μl)
Vial 4
Purified apoB standard
To ensure total recovery of stated quantity, vials have been overfilled.
STORAGE:
Shipped at ambient temperature. On arrival box 1 should be stored
refrigerated at 4-8°C and box 2 should be stored frozen at -20°C.

GeneralIntended use: For quantitative determination of human Apolipoprotein B (apoB) in serum/plasma samples or cell culture supernatants. This kit is specific for the detection of apoB100 anddoes not recognize apoB48. The mAbs recognize apoB100 in its form as VLDL/LDL associatedprotein, whereas purified delipified apoB is poorly recognized.
Please note that wash-, block- and incubation buffers should contain detergent. Tween 20, Triton
X-100 or NP40 can be used at a concentration of 0.05-0.5%. In block and incubation buffers it isrecommended to use 0.1% BSA.
Serum/plasma samples: To prevent interference by different LDL-particle sizes, serum/plasma
samples should be treated with Triton X-100. Dilute samples 2x with 1% Triton X followed byvortex for 5 seconds. Triton X-treatment is not necessary for the apoB standard or for cell line
produced samples. When analyzing human serum/plasma samples it is recommended to use Apo
ELISA buffer (product code: 3652-M2) for dilution of samples, standard and detection antibody.
The buffer prevents false positive read-outs which may be caused by interference of heterophilic
antibodies commonly found in human plasma and serum. The Apo ELISA buffer has been validated using serum/plasma from normal healthy human blood donors. Please note that heterophilic
antibody interference in samples from human subjects with various diseases or other conditions
has not been assessed. Freezing and thawing serum/plasma will reduce the recognition of apoB bythese antibodies. It is recommended to dilute serum/plasma samples 5,000x to 8,000x. Please see
dilution guidelines at https://www.mabtech.com/knowledge-center/apodilution.
Reagents: Antibodies are supplied in sterile-filtered (0.2 μm) PBS with sodium azide (0.02%).
Streptavidin-ALP is supplied in 0.1 M Tris buffer with 0.002% Kathon CG.
Standard range: 8-800 ng/ml
Standard calibration: No international standard exists for calibration.

OD 405 nm

1010,10,01

110100Concentration of apoB standard (ng/ml)

1000

Guidelines for Human Apolipoprotein B ELISA
Day 1 1.

Coat a high protein binding ELISA plate with mAb LDL 20/17, diluted to 2 μg/ml
in PBS, pH 7.4, by adding 100 μl/well. Incubate overnight at 4-8°C.

Day 2 2.

Wash twice with PBS (200 μl/well).

3.

Block plate by adding 200 μl/well of PBS with 0.05% Tween 20 containing 0.1%
BSA (incubation buffer). Incubate for 1 hour at room temperature.

4.

Wash five times with PBS containing 0.05% Tween.

5.

The apoB standard is supplied as purified LDL stabilised by 50% glycerol. The concentration is 125 μg/ml. For the test, prepare dilutions of the stock using the standard range as a guideline.

6.

Add 100 μl/well of samples or standards diluted in incubation buffer or Apo ELISA
buffer for serum/plasma samples and incubate for 1 to 2 hours at room temperature.
Please note the special considerations for serum/plasma samples described above.
Dilution recommendations for serum/plasma samples can be found at https://www.
mabtech.com/knowledge-center/apodilution.

7.

Wash as in step 4.

8.

Add 100 μl/well of mAb LDL 11-biotin at 1 μg/ml in incubation buffer or Apo
ELISA buffer for serum/plasma samples. Incubate for 1 hour at room temperature.

9.

Wash as in step 4.

10. Add 100 μl/well of Streptavidin-ALP diluted 1:1000 in incubation buffer. Incubate
for 1 hour at room temperature.
11. Wash as in step 4.
12. Add 100 μl/well of appropriate substrate solution e.g. p-nitrophenyl-phosphate(pNPP), available from Mabtech product code 3652-P10.
13. Measure the optical density (405 nm for pNPP) in an ELISA reader after suitabledeveloping time.

Updated on 2020/03/31

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Developed and manufactured by MABTECH AB, Sweden,
whose quality management system complies with thestandards ISO 9001:2015 & ISO 13485:2016.

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The products are for research use only.
MABTECH shall not be liable for the use or handling of the product or for consequential,special, indirect or incidental damages there from.
Mabtech AB (Head Office)SwedenTel: +46 8 716 27 00
mabtech@mabtech.com

www.mabtech.com

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USA
Tel: +1 513 871-4500
mabtech.usa@mabtech.com