- MIA1402 - Invitrogen Antibodies APOA1 antibody

Submitted references Serum HDL cholesterol uptake capacity in subjects from the MASHAD cohort study: Its value in determining the risk of cardiovascular endpoints.

Aghasizadeh M, Samadi S, Sahebkar A, Miri-Moghaddam E, Esmaily H, Souktanloo M, Avan A, Mansoori A, Ferns GA, Kazemi T, Ghayour-Mobarhan M
Journal of clinical laboratory analysis 2021 Jun;35(6):e23770

Association of ANGPTL3 polymorphisms with high-density lipoprotein cholesterol uptake capacity in patients with cardiovascular disease.

Aghasizadeh M, Nosrati M, Saberi-Karimian M, Safarian H, Assadian P, Akbarpour E, Sahebkar A, Avan A, Ferns GA, Kazemi T, Miri-Moghaddam E, Ghayour-Mobarhan M
Journal of clinical laboratory analysis 2021 Dec;35(12):e23980

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of Apolipoprotein A-1 was performed by loading the indicated amounts of a recombinant human Apo A-1-hIgG1Fc fusion protein (Product # 10686-H02H-50) or a recombinant mouse Apo A-1-hIgG1Fc fusion protein (Product # 50918-M02H-50), and 10 µL of PageRuler Prestained Protein Ladder (Product # 26616) per well onto a Novex® 4-20% Tris-Glycine polyacrylamide gel (Product # WT4202BOX ). Proteins were transferred to a Nitrocellulose Membrane (Product # 88014) using the G2 Fast Blotter (Product # 62288), and blocked with 5% milk in TBST for at least 1 hour at room temperature. Apo A-1 was detected at ~50 kD using an Apolipoprotein A-1 monoclonal antibody (Product # MIA1402) at a dilution of 1:1000 in blocking buffer overnight at 4C on a rocking platform, followed by an HRP-conjugated goat anti-mouse IgG Fc-specific secondary antibody (Product # 31439) at a dilution of 1:40,000 for at least 30 minutes at room temperature. Chemiluminescent detection was performed using SuperSignal West Dura (Product # 34075).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of Apolipoprotein A-1 was performed by loading the indicated amounts of a recombinant human Apo A-1-hIgG1Fc fusion protein (Product # 10686-H02H-50) or a recombinant mouse Apo A-1-hIgG1Fc fusion protein (Product # 50918-M02H-50), and 10 µL of PageRuler Prestained Protein Ladder (Product # 26616) per well onto a Novex® 4-20% Tris-Glycine polyacrylamide gel (Product # WT4202BOX ). Proteins were transferred to a Nitrocellulose Membrane (Product # 88014) using the G2 Fast Blotter (Product # 62288), and blocked with 5% milk in TBST for at least 1 hour at room temperature. Apo A-1 was detected at ~50 kD using an Apolipoprotein A-1 monoclonal antibody (Product # MIA1402) at a dilution of 1:1000 in blocking buffer overnight at 4C on a rocking platform, followed by an HRP-conjugated goat anti-mouse IgG Fc-specific secondary antibody (Product # 31439) at a dilution of 1:40,000 for at least 30 minutes at room temperature. Chemiluminescent detection was performed using SuperSignal West Dura (Product # 34075).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig 1: Western blot analysis was performed on tissue extract (30 µg lysate) of Mouse Liver (Lane 1). Fig 2: Likewise, Western blot analysis was performed on samples of 3 µL serum (Lane 1), 5 µL serum (Lane 2), 5 µL plasma (Lane 3), all of which were obtained from a 50 year old male Alzheimer's patient. The blots were probed with Anti-Apolipoprotein A-1 Mouse Monoclonal Antibody (Product # MIA1402, 1:500-1:2000 dilution) and detected by chemiluminescence Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.4 µg/mL, 1:2500 dilution). A 28 kDa band corresponding to Apolipoprotein A-1 was observed across the tissues and samples tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 12 % Bis-Tris gel (Product # NP0342BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5 % skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Sandwich ELISA of Apolipoprotein A-1 was performed by coating wells of a 96-well plate with 100 µL of an Apo A-1 Recombinant Rabbit Polyclonal Antibody (Product # 710263) at a concentration of 1 µg/mL in carbonate/bicarbonate buffer (Product # 28382) overnight at 4C. Wells were blocked with 150 µL of StartingBlock T20 (TBS) Blocking Buffer (Product # 37543) for 30 minutes, and 80 µL of recombinant human Apo A-1 (Product # 10686-H02H-50) standards ranging from 1.6-1000 ng/mL (A) or 100 µL of diluted normal human serum or diluted serum from a patient with dyslipidemia (B) were incubated for 1 hour at room temperature. The plate was washed with 1X TBST (Product # 28360), and 100 µL per well of an Apo A-1 Mouse Monoclonal Antibody (Product # MIA1402) was added to each well for 1 hour at room temperature. The plate was washed, and 100 µL per well of a biotinylated Rabbit anti-Mouse IgG Superclonal secondary antibody (Product # A27024) was incubated for 1 hour, followed by Streptavidin-HRP (Product # 21126) for 30 minutes. Detection was performed using 1-Step Ultra TMB Substrate (Product # 34028), followed by Stop Solution (Product # N600). Absorbances were read on a spectrophotometer at 450-550 nm.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescence analysis of Apolipoprotein A1 was performed 70% confluent log phase Hep G2 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 2% BSA for 1 hour at room temperature. The cells were labeled with Apolipoprotein A1 (311) Mouse Monoclonal Antibody (Product # MIA1402) at 2 µg/mL in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A28175) a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing cytoplasmic localization. Panel e shows the no primary antibody control. The images were captured at 60X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of Apolipoprotein A-1 showing staining in the cytoplasm of paraffin-embedded human liver tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a Apolipoprotein A-1 Mouse Monoclonal Antibody (Product # MIA1402) diluted in 3% BSA-PBS at a dilution of 1:100 for 1 hour at 37ºC in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow cytometry analysis of Apolipoprotein A-1 was done on Hep G2 cells. Cells were fixed with 70% ethanol for 10 minutes, permeabilized with 0.25% Triton™ X-100 for 20 minutes, and blocked with 5% BSA for 30 minutes at room temperature. Cells were labeled with Apolipoprotein A-1 Mouse Monoclonal Antibody (MIA1402, red histogram) or with mouse isotype control (pink histogram) at 3-5 ug/million cells in 2.5% BSA. After incubation at room temperature for 2 hours, the cells were labeled with Alexa Fluor® 488 Rabbit Anti-Mouse Secondary Antibody (A11059) at a dilution of 1:400 for 30 minutes at room temperature. The representative 10,000 cells were acquired and analyzed for each sample using an Attune® Acoustic Focusing Cytometer. The purple histogram represents unstained control cells and the green histogram represents no-primary-antibody control..

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Sandwich ELISA of Apolipoprotein A-1 was performed by coating wells of a 96-well plate with 100 µL of an Apo A-1 Recombinant Rabbit Polyclonal Antibody (Product # 710263) diluted to a concentration of 1 µg/mL in carbonate/bicarbonate buffer (Product # 28382) overnight at 4C. Wells were blocked with 150 µL of StartingBlock T20 (TBS) Blocking Buffer (Product # 37543) for 30 minutes, and 80 µL of recombinant human Apo A-1 (Product # 10686-H02H-50) or recombinant mouse Apo A-1 (Product # 50918-M02H-50) was added to the plate at concentrations ranging from 1.6-1000 ng/mL and incubated for 1 hour at room temperature. The plate was washed with 1X TBST (Product # 28360), and 100 µL per well of an Apo A-1 Mouse Monoclonal Antibody (Product # MIA1402) was added to each well for 1 hour at room temperature. The plate was washed, and 100 µL per well of an HRP-conjugated Rabbit anti-Mouse IgG cross-adsorbed secondary antibody (Product # 31452) was incubated for 30 minutes at room temperature. Detection was performed using 1-Step Ultra TMB Substrate (Product # 34028), followed by Stop Solution (Product # N600). Absorbances were read on a spectrophotometer at 450-550 nm.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Sandwich ELISA of Apolipoprotein A-1 was performed by coating wells of a 96-well plate with 100 µL of an Apo A-1 rabbit monoclonal antibody (Product # 701239) at a concentration of 1 µg/mL in carbonate/bicarbonate buffer (Product # 28382) overnight at 4C. Wells were blocked with 150 µL of StartingBlock T20 (TBS) Blocking Buffer (Product # 37543) for 30 minutes, and 80 µL of recombinant human Apo A-1 (Product # 10686-H02H-50) standards ranging from 1.6-1000 ng/mL (A) or 100 µL of diluted normal human serum or diluted serum from a patient with dyslipidemia (B) were incubated for 1 hour at room temperature. The plate was washed with 1X TBST (Product # 28360), and 100 µL per well of an Apo A-1 mouse monoclonal antibody (Product # MIA1402) was added to each well for 1 hour at room temperature. The plate was washed, and 100 µL per well of a biotinylated rabbit anti-mouse IgG Superclonal? secondary antibody (Product # A27024) was incubated for 1 hour, followed by Streptavidin-HRP (Product # 21126) for 30 minutes. Detection was performed using 1-Step Ultra TMB Substrate (Product # 34028), followed by Stop Solution (Product # N600). Absorbances were read on a spectrophotometer at 450-550 nm.

MIA1402

Antibody data