Recombinant Anti-Lin28B antibody [EPR18717] KO Tested (ab191881)

Product name

Anti-Lin28B antibody [EPR18717]
See all Lin28B primary antibodies
Description

Rabbit monoclonal [EPR18717] to Lin28B
Host species

Rabbit

Tested Applications & Species

Application	Species
Flow Cyt	Human
ICC/IF	Human
IP	Human
WB	Human Recombinant fragment

See all applications and species data

Immunogen

Recombinant full length protein. This information is proprietary to Abcam and/or its suppliers.
Positive control
- WB: Recombinant full length Human Lin28B protein; Human placenta and testis lysates; K562, HepG2, HEK-293T and NCCIT whole cell lysates. ICC/IF: JAR and K562 cells. ICC/IF KO: HEK293 cells (HEK293-Lin28B KO used as a negative cell line) Flow Cyt: K562 cells. IP: K562 whole cell lysate.
General notes
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
For more information see here.
Our RabMAb^® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb^® patents.
The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing the problem with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation.

One factor contributing to the crisis is the use of antibodies that are not suitable. This can lead to misleading results and the use of incorrect data informing project assumptions and direction. To help address this challenge, we have introduced an application and species grid on our primary antibody datasheets to make it easy to simplify identification of the right antibody for your needs.

Learn more here.

Form

Liquid
Storage instructions

Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
Storage buffer

pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Concentration information loading...
Purity

Protein A purified
Clonality

Monoclonal
Clone number

EPR18717
Isotype

IgG
Research areas

Alternative Versions
- Anti-Lin28B antibody [EPR18717] - BSA and Azide free (ab240326)
Compatible Secondaries
- Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077)
- Goat Anti-Rabbit IgG H&L (HRP) (ab97051)
Conjugation kits
- PE / R-Phycoerythrin Conjugation Kit - Lightning-Link® (ab102918)
- APC Conjugation Kit - Lightning-Link® (ab201807)
Isotype control
- Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)
KO cell lines
- Human LIN28B knockout HEK-293T cell line (ab265066)
KO cell pellets
- Human LIN28B knockout HEK293T cell pellet (ab278864)
Recombinant Protein
- Recombinant Human Lin28B protein (ab134596)

The Abpromise guarantee

Our Abpromise guarantee covers the use of ab191881 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Guaranteed

Tested applications are guaranteed to work and covered by our Abpromise guarantee.

Predicted

Predicted to work for this combination of applications and species but not guaranteed.

Incompatible

Does not work for this combination of applications and species.

Application	Species
Flow Cyt	Human
ICC/IF	Human
IHC-P	Mouse Rat Human
IP	Human
WB	Human Recombinant fragment

Application	Abreviews	Notes
WB		1/2000. Detects a band of approximately 30 kDa (predicted molecular weight: 27 kDa).
ICC/IF		Use a concentration of 1 - 10 µg/ml.
Flow Cyt		1/150.
IP		1/50.

Notes
WB 1/2000. Detects a band of approximately 30 kDa (predicted molecular weight: 27 kDa).
ICC/IF Use a concentration of 1 - 10 µg/ml.
Flow Cyt 1/150.
IP 1/50.

Function

Acts as a suppressor of microRNA (miRNA) biogenesis by specifically binding the precursor let-7 (pre-let-7), a miRNA precursor. Acts by binding pre-let-7 and recruiting ZCCHC11/TUT4 uridylyltransferase, leading to the terminal uridylation of pre-let-7. Uridylated pre-let-7 miRNAs fail to be processed by Dicer and undergo degradation. Specifically recognizes the 5'-GGAG-3' motif in the terminal loop of pre-let-7. Also recognizes and binds non pre-let-7 pre-miRNAs that contain the 5'-GGAG-3' motif in the terminal loop, leading to their terminal uridylation and subsequent degradation. Mediates MYC-mediated let-7 repression. Isoform 1, when overexpressed, stimulates growth of the breast adenocarcinoma cell line MCF-7. Isoform 2 has no effect on cell growth.
Tissue specificity

High expression in testis, fetal liver, placenta and in hepatocellular carcinoma (HCC). Isoform 1 is only detected in moderately and poorly differentiated HCC tissues and placenta (at protein level). Isoform 2 is detected in fetal liver, non-tumor liver tissues, as well as well-differentiated tumor tissues (at protein level).
Sequence similarities

Belongs to the lin-28 family.
Contains 2 CCHC-type zinc fingers.
Contains 1 CSD (cold-shock) domain.
Cellular localization

Cytoplasm. Nucleus. Predominantly cytoplasmic at G1 phase, accumulates in the nucleus in S and G2 phases. The frequency of nuclear localization in S and in G2 phases is 60% and 30%, respectively.
Target information above from: UniProt accession Q6ZN17 The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010) .

Information by UniProt
Database links
- Entrez Gene: 389421 Human
- Omim: 611044 Human
- SwissProt: Q6ZN17 Human
- Unigene: 23616 Human
Alternative names
- CSDD 2 antibody
- CSDD2 antibody
- FLJ16517 antibody
- Lin 28 homolog B (C. elegans) antibody
- Lin 28.2 antibody
- Lin-28B antibody
- LIN28B antibody
- LN28B_HUMAN antibody
- Protein lin-28 homolog B antibody
see all

Western blot - Anti-Lin28B antibody [EPR18717] (ab191881)

All lanes : Anti-Lin28B antibody [EPR18717] (ab191881) at 1/2000 dilution

Lane 1 : Wild-type HEK293 cell lysate
Lane 2 : LIN28B knockout HEK293 cell lysate
Lane 3 : HepG2 cell lysate
Lane 4 : K562 cell lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 27 kDa
Observed band size: 35 kDa why is the actual band size different from the predicted?

Lanes 1 - 4: Merged signal (red and green). Green - ab191881 observed at 35 kDa. Red - loading control ab7291 (Mouse anti-Alpha Tubulin [DM1A]) observed at 55 kDa.

ab191881 was shown to react with Lin28B in wild-type HEK-293 cells in Western blot with loss of signal observed in LIN28B knockout sample. Wild-type HEK-293 and LIN28B knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween^®) before incubation with ab191881 and ab7291 (Mouse anti-Alpha Tubulin [DM1A]) overnight at 4°C at a 1 in 2000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.
Immunocytochemistry/ Immunofluorescence - Anti-Lin28B antibody [EPR18717] (ab191881)

ab191881 staining Lin28B in wild-type HEK293 cells (top panel) and Lin28B knockout HEK293 cells (bottom panel). The cells were fixed with 4% paraformaldehyde (10 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab191881 at 10µg/ml and ab7291 (Mouse monoclonal to alpha Tubulin) at 1/1000 dilution overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to rabbit IgG (Alexa Fluor® 488) (ab150081) at 2 μg/ml (shown in green) and a goat secondary antibody to mouse IgG (Alexa Fluor® 594) (ab150120) at 2 μg/ml (shown in pseudo color red). Nuclear DNA was labelled in blue with DAPI.

Image was taken with a high-content analysis system (Perkin Elmer, Operetta CLS™).
Western blot - Anti-Lin28B antibody [EPR18717] (ab191881)

All lanes : Anti-Lin28B antibody [EPR18717] (ab191881) at 1/1000 dilution

Lane 1 : Wild-type HEK293T cell lysate
Lane 2 : LIN28B knockout HEK293T cell lysate
Lane 3 : HepG2 cell lysate
Lane 4 : SW480 cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/10000 dilution

Predicted band size: 27 kDa
Observed band size: 36 kDa why is the actual band size different from the predicted?

Lanes 1-4: Merged signal (red and green). Green - ab191881 observed at 36 kDa. Red - loading control ab7291 observed at 50 kDa.

ab191881 Anti-Lin28B antibody [EPR18717] was shown to specifically react with Lin28B in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab265066 (knockout cell lysate ab257504) was used. Wild-type and Lin28B knockout samples were subjected to SDS-PAGE. ab191881 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
Western blot - Anti-Lin28B antibody [EPR18717] (ab191881)

Anti-Lin28B antibody [EPR18717] (ab191881) at 1/2000 dilution + Recombinant full length Human Lin28B protein at 0.01 µg

Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution

Predicted band size: 27 kDa

Exposure time: 1 second

Blocking/Dilution buffer: 5% NFDM/TBST.
Recombinant Human Lin28B protein contains aa1-250 with His-Tag®.
Western blot - Anti-Lin28B antibody [EPR18717] (ab191881)

Anti-Lin28B antibody [EPR18717] (ab191881) at 1/2000 dilution + Human placenta lysate at 20 µg

Secondary
Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/10000 dilution

Predicted band size: 27 kDa
Observed band size: 30 kDa why is the actual band size different from the predicted?

Exposure time: 3 minutes

Blocking/Dilution buffer: 5% NFDM/TBST.
Western blot - Anti-Lin28B antibody [EPR18717] (ab191881)

Anti-Lin28B antibody [EPR18717] (ab191881) at 1/2000 dilution + Human testis lysate at 10 µg

Secondary
Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/10000 dilution

Predicted band size: 27 kDa
Observed band size: 30 kDa why is the actual band size different from the predicted?

Exposure time: 3 minutes

Blocking/Dilution buffer: 5% NFDM/TBST.
Western blot - Anti-Lin28B antibody [EPR18717] (ab191881)

All lanes : Anti-Lin28B antibody [EPR18717] (ab191881) at 1/2000 dilution

Lane 1 : K562 (Human chronic myelogenous leukemia cells from bone marrow) whole cell lysate
Lane 2 : HepG2 (Human liver hepatocellular carcinoma) whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution

Predicted band size: 27 kDa
Observed band size: 30 kDa why is the actual band size different from the predicted?

Exposure time: 5 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.
Western blot - Anti-Lin28B antibody [EPR18717] (ab191881)

Anti-Lin28B antibody [EPR18717] (ab191881) at 1/2000 dilution + NCCIT (Human pluripotent embryonic carcinoma) whole cell lysate at 20 µg

Secondary
Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution

Predicted band size: 27 kDa
Observed band size: 30 kDa why is the actual band size different from the predicted?

Exposure time: 15 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.
Immunocytochemistry/ Immunofluorescence - Anti-Lin28B antibody [EPR18717] (ab191881)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized JAR (Human placenta choriocarcinoma cell line) cells labeling Lin28B with ab191881 at 1/1000 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasm and nuclear staining on JAR cells. The nuclear counter stain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).
The negative controls are as follows:
-ve control 1: ab191881 at 1/1000 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
-ve control 2: ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.
Immunocytochemistry/ Immunofluorescence - Anti-Lin28B antibody [EPR18717] (ab191881)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized K562 (Human chronic myelogenous leukemia cells from bone marrow) cells labeling Lin28B with ab191881 at 1/1000 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing cytoplasm and nuclear staining on K562 cells. The nuclear counter stain is DAPI (blue). Tubulin is detected with ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution and ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution (red).
The negative controls are as follows:
-ve control 1: ab191881 at 1/1000 dilution followed by ab150120 (AlexaFluor®594 Goat anti-Mouse secondary) at 1/1000 dilution.
-ve control 2: ab7291 (anti-Tubulin mouse mAb) at 1/1000 dilution followed by ab150077 (Alexa Fluor®488 Goat Anti-Rabbit IgG H&L) at 1/1000 dilution.
Flow Cytometry - Anti-Lin28B antibody [EPR18717] (ab191881)

Flow cytometric analysis of 4% paraformaldehyde-fixed K562 (Human chronic myelogenous leukemia cells from bone marrow) cells labeling Lin28B with ab191881 at 1/150 dilution (red) compared with a rabbit monoclonal IgG isotype control (ab172730; black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody; blue). Goat anti rabbit IgG (FITC) at 1/500 dilution was used as the secondary antibody.
Immunoprecipitation - Anti-Lin28B antibody [EPR18717] (ab191881)

Lin28B was immunoprecipitated from 1mg of K562 (Human chronic myelogenous leukemia cells from bone marrow) whole cell lysate with ab191881 at 1/50 dilution. Western blot was performed from the immunoprecipitate using ab191881 at 1/1000 dilution. Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG, was used as secondary antibody at 1/1500 dilution.
Lane 1: K562 whole cell lysate 10ug (Input).
Lane 2: ab191881 IP in K562 whole cell lysate.
Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab191881 in K562 whole cell lysate.
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
Exposure time: 5 seconds.
Anti-Lin28B antibody [EPR18717] (ab191881)

Flow cytometry protocols
Immunoprecipitation protocols
Immunocytochemistry & immunofluorescence protocols
Western blot protocols

Click here to view the general protocols

SDS download

Country/Region

Language
Datasheet download

Download

Publishing research using ab191881? Please let us know so that we can cite the reference in this datasheet.

ab191881 has been referenced in 4 publications.

Mizushima E et al. Osteosarcoma-initiating cells show high aerobic glycolysis and attenuation of oxidative phosphorylation mediated by LIN28B. Cancer Sci 111:36-46 (2020). PubMed: 31705593
Vishnubalaji R et al. Neoplastic Transformation of Human Mesenchymal Stromal Cells Mediated via LIN28B. Sci Rep 9:8101 (2019). PubMed: 31147574
Wang X et al. RNA binding protein Lin28B confers gastric cancer cells stemness via directly binding to NRP-1. Biomed Pharmacother 104:383-389 (2018). PubMed: 29787985
Zhao G et al. Placental exosome-mediated Bta-miR-499-Lin28B/let-7 axis regulates inflammatory bias during early pregnancy. Cell Death Dis 9:704 (2018). PubMed: 29899331

Key features and details

Overview

Product name

Description

Host species

Tested Applications & Species

Immunogen

Positive control

General notes

Properties

Form

Storage instructions

Storage buffer

Purity

Clonality

Clone number

Isotype

Research areas

Associated products

Alternative Versions

Compatible Secondaries

Conjugation kits

Isotype control

KO cell lines

KO cell pellets

Recombinant Protein

Applications

The Abpromise guarantee

Target

Function

Tissue specificity

Sequence similarities

Cellular localization

Database links

Alternative names

Images

Protocols

Datasheets and documents

SDS download

Datasheet download

Certificate of Compliance

References (4)

Customer reviews and Q&As