A validated antibody toolbox for ALS research
- Structural Genomics Consortium, Montreal Neurological Institute-Hospital (The Neuro), McGill University, Canada
- Neurodegenerative Diseases Research Group, Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital (The Neuro), McGill University, Canada
- NIHR Respiratory Biomedical Research Centre, Department of Respiratory Sciences, University of Leicester, United Kingdom
- Early Drug Discovery Unit (EDDU), Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital (The Neuro), McGill University, Canada
- Department of Genetics, Stanford University School of Medicine, United States
- Structural Genomics Consortium, Nuffield Department of Medicine, University of Oxford, United Kingdom
- Centre for Human Genetics and Centre for Medicines Discovery, Nuffield Department of Medicine, University of Oxford, United Kingdom
- Nuffield Department of Medicine, University of Oxford, United Kingdom
- Structural Genomics Consortium, Department of Medicine, Karolinska Institute, Sweden
- Neuroimmunology Unit, Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital (The Neuro), McGill University, Canada
- School of Pharmaceutical Sciences, Faculty of Medicine, University of Ottawa, Canada
Figures
Figure 1 with 5 supplements
Summary of antibody performance for ALS-RAP proteins.
(A) Overview of the identification of high-quality antibodies targeting ALS-RAP proteins. (B) Percentage of ALS-RAP targets covered by at least one specific renewable (monoclonal or recombinant) antibody for the indicated application. (C) Percentage of tested antibodies that met specificity criteria in each indicated application.
Figure 1—figure supplement 1
Characterization of ALS-RAP targets.
Number of PubMed entries related to ALS-RAP genes. Black points represent the number of entries retrieved using the search term “gene name + ALS,” while gray points represent entries retrieved using the gene name alone. The x-axis indicates the number of PubMed entries, and the y-axis lists the gene names.
Figure 1—figure supplement 2
ALS-RAP RNA expression across brain cell types.
Heatmap showing normalized RNA expression values from the Human Protein Atlas (HPA) single-nuclei brain dataset. Expression is given as nTPM values on a scale from 0 to 500. Genes denoted with an asterisk (*) are classified as “barely detectable” but show substantial expression in a single-cell type.
Figure 1—figure supplement 3
Protein-level characterization of iPSC-derived neurological cells.
(A) Table listing cell type–specific marker proteins. (B) WB analysis of marker proteins using antibodies against the indicated proteins. A Ponceau-stained membrane is shown to assess protein loading and transfer efficiency. Lysates were prepared from the indicated human iPSC-derived cell types or fetal astrocyte culture. Vertical lines between the motor neuron and DA neuron lanes and between the DA neuron and microglia lanes indicate that WB were spliced to remove additional conditions not relevant to this study. DA, dopaminergic; NPC, neuronal progenitor cells.
Figure 1—figure supplement 4
Quality control and marker validation of iPSC-derived microglia.
(A) Immunofluorescence staining showing expression of IBA1 and PU.1 in iMGs, with no detectable signal in undifferentiated iPSCs. (B) Representative flow cytometry plots demonstrating expression of CD45 and CD11b in iMGs but not in iPSCs. (C) Quantification of the percentage of CD45-positive, CD11b-positive, and double-positive cells in iPSCs and iMGs, as measured by flow cytometry (N=4). Statistical analysis was performed using two-way ANOVA with Bonferroni post hoc test. (D) RT-qPCR expression levels of canonical microglial markers in iMGs compared to iPSCs (N=3). Statistical analysis was performed using Student’s t-test. iMG, iPSC-derived microglia; iPSC, induced pluripotent stem cell. *p≤0.05, **p≤0.01, ***p≤0.001, ****p≤0.0001. Scale bar, 50 µm.
Figure 1—figure supplement 5
STRING analysis of known physical interactions between ALS-RAP proteins.
Figure 2
YCharOS antibody characterization workflow for Galectin-1 (LGALS1).
Ten Galectin-1 antibodies were evaluated side-by-side across three applications using HAP1 WT and LGALS1 KO cells. (A) Whole-cell protein lysates from HAP1 WT and LGALS1 KO cells (20 µg per lane) were analyzed in WB using the indicated Galectin-1 antibodies. Ponceau-stained membranes are shown to assess protein loading and transfer efficiency. (B) Whole-cell protein lysates (10 µg) and conditioned media (10 µg) from HAP1 WT and LGALS1 KO cells were analyzed in WB using antibody 12936, as in (A), to detect intracellular and secreted Galectin-1. (C) Immunoprecipitation was performed from HAP1 WT cell lysates using the indicated Galectin-1 antibodies. Starting material (SM), unbound fractions (UB), and whole immunoprecipitates (IP) were analyzed by WB. Anti-Galectin-1 antibody A21218 was used as a primary antibody. (D) HAP1 WT and LGALS1 KO cells were differentially labeled with green and far-red fluorescent dyes, respectively, mixed at a 1:1 ratio, and plated in optically clear 96-well plates. Cells were stained with the indicated Galectin-1 antibodies, followed by Coralite 555–conjugated secondary antibodies and DAPI. Images were acquired in the blue (DAPI), green (WT), red (antibody staining), and far-red (KO) channels. Representative grayscale images of the blue and red channels are shown; WT and KO cells are outlined with green and magenta dashed lines, respectively. Scale bar, 10 µm. Recombinant antibodies are denoted by double asterisks (**) and monoclonal antibodies by single asterisks (*) after the catalogue numbers; polyclonal antibodies are unmarked. Antibody screening was performed in two independent experiments; representative data from one experiment are shown. Galectin-1 antibody characterization raw datasets are available at https://doi.org/10.5281/zenodo.21538630.
Figure 3
Accessing curated antibody characterization data through the OGA website.
YCharOS antibody characterization data for ALS-RAP proteins are accessible through the Only Good Antibodies (OGA) website (https://onlygoodantibodies.co.uk/). The OGA homepage provides a searchable interface for the 33 ALS-RAP proteins, together with 35 additional neurological disease-associated proteins evaluated by YCharOS, and includes links to key OGA publications, data interpretation guidelines, antibody characterization protocols, and a portal for nominating new targets. (A) Representative results page generated by searching LGALS1, showing filters for host species, clonality, recombinant format, and OGA-recommended applications; the WB filter is applied. A schematic panel at the top illustrates ideal antibody performance across applications, while antibodies matching the selected filters are displayed in randomized order. (B) Number of antibodies returned following application of the selected filters.
Figure 4
Detection of 13 ALS-RAP proteins across five human-derived neurological cell types and three primary cell types.
WB showing detection of 13 ALS-RAP proteins in eight lysates from the indicated five iPSC-derived cell types and three primary cell types. For each protein, two independent experiments are shown: chemiluminescence-based WB (left), performed using a defined lysate batch across all corresponding blots, and fluorescence-based WB (right), performed using a second independent lysate batch. Proteins are listed in alphabetical order. Vertical lines between the motor neuron and DA neuron lanes indicate that the blots were spliced to remove additional conditions not relevant to this study. Lane identities: 1, motor neurons; 2, DA neurons; 3, oligodendrocytes; 4, astrocytes; 5, microglia; 6, fetal astrocytes; 7, fetal microglia; 8, MDMs. DA, dopaminergic; MDM, monocyte-derived macrophages. Raw WB data and supporting working sheet for the ALS-RAP protein expression profiling across neurological cell types by chemiluminescent WB are available at https://doi.org/10.5281/zenodo.18484385. Raw WB data and supporting working sheet for the ALS-RAP protein expression profiling across neurological cell types by fluorescent WB are available at https://doi.org/10.5281/zenodo.18484483.
Figure 5
Detection of 10 ALS-RAP proteins across five human-derived neurological cell types and two primary cell types.
WB showing detection of 10 ALS-RAP proteins in 7 lysates from the indicated five iPSC-derived cell types and 2 primary cell types. Fetal astrocyte lysate was not available for either chemiluminescence-based WB (left) and fluorescence-based WB (right). Proteins are listed in alphabetical order. Lane identities: 1, motor neurons; 2, DA neurons; 3, oligodendrocytes; 4, astrocytes; 5, microglia; 7, fetal microglia; 8, MDMs. DA, dopaminergic; MDM, monocyte-derived macrophages. Raw WB data and supporting working sheet for the ALS-RAP protein expression profiling across neurological cell types by chemiluminescent WB are available at https://doi.org/10.5281/zenodo.18484385. Raw WB data and supporting working sheet for the ALS-RAP protein expression profiling across neurological cell types by fluorescent WB are available at https://doi.org/10.5281/zenodo.18484483.
Figure 6
Detection of seven ALS-RAP proteins across five human-derived neurological cell types and primary cell types.
(A) Chemiluminescence-based WB (left) showing detection of seven ALS-RAP proteins in seven lysates from the indicated five iPSC-derived cell types and two primary cell types. Fetal astrocyte lysate was not available for this WB. Fluorescence-based WB (right) showing detection of seven ALS-RAP proteins in eight lysates from the indicated five iPSC-derived cell types and three primary cell types. A representative Ponceau-stained membrane and total protein stain of all eight samples are shown to exemplify protein loading and transfer efficiency. Lane identities: 1, motor neurons; 2, DA neurons; 3, oligodendrocytes; 4, astrocytes; 5, microglia; 6, fetal astrocytes; 7, fetal microglia; 8, MDMs. DA, dopaminergic; MDM, monocyte-derived macrophages. (B) Summary of 16 targets for which WB data and Human Protein Atlas RNA expression profiles are concordant, grouped by expression category. Raw WB data and supporting working sheet for the ALS-RAP protein expression profiling across neurological cell types by chemiluminescent WB are available at https://doi.org/10.5281/zenodo.18484385. Raw WB data and supporting working sheet for the ALS-RAP protein expression profiling across neurological cell types by fluorescent WB are available at https://doi.org/10.5281/zenodo.18484483.
Tables
Table 1
ALS-associated genes selected for the ALS-RAP study.
| Gene name | UniProt ID | Effect size (variants; effect size) | ClinGen curated description | PubMed entries (gene name +ALS) | PubMed entries (gene name) |
|---|---|---|---|---|---|
| SOD1 | P00441 | Rare; high | Definitive | 4679 | 10,855 |
| TARDBP | Q13148 | Rare; high | Definitive | 2846 | 5061 |
| C9orf72 | Q96LT7 | Rare; high | Definitive | 1952 | 2757 |
| FUS | P35637 | Rare; high | Definitive | 1332 | 5694 |
| VCP | P55072 | Rare; high | Definitive | 267 | 2354 |
| OPTN | Q96CV9 | Rare; high | Definitive | 206 | 1640 |
| SQSTM1 | Q13501 | Rare; intermediate | Moderate | 183 | 3848 |
| TBK1 | Q9UHD2 | Rare; high | Definitive | 178 | 2749 |
| VAPB | O95292 | Rare; intermediate | Definitive | 169 | 417 |
| ATXN2 | Q99700 | Rare; intermediate | Definitive | 163 | 437 |
| UBQLN2 | Q9UHD9 | Rare; high | Definitive | 139 | 206 |
| ALS2 | Q96Q42 | Rare; intermediate | Definitive | 127 | 250 |
| ANG | P03950 | Rare; intermediate | Limited | 99 | 1592 |
| SETX | Q7Z333 | Rare; intermediate | Not available | 93 | 257 |
| CHCHD10 | Q8WYQ3 | Rare; intermediate | Moderate | 93 | 174 |
| HNRNPA1 | P09651 | Rare; unknown | Not available | 91 | 574 |
| PFN1 | P07737 | Rare; intermediate | Definitive | 79 | 356 |
| TAF15 | Q92804 | Rare; intermediate | Limited | 66 | 251 |
| NEK1 | Q96PY6 | Rare; high | Definitive | 64 | 192 |
| MATR3 | P43243 | Rare; unknown | Definitive | 61 | 162 |
| KIF5A | Q12840 | Rare; intermediate | Definitive | 61 | 259 |
| CHMP2B | Q9UQN3 | Rare; intermediate | Definitive | 48 | 225 |
| TIA1 | P31483 | Rare; intermediate | Limited | 47 | 1146 |
| ANXA11 | P50995 | Rare; intermediate | Definitive | 47 | 124 |
| CCNF | P41002 | Rare; small | Limited | 41 | 184 |
| FIG4 | Q92562 | Rare; intermediate | Limited | 40 | 170 |
| HNRNPA2B1 | P22626 | Rare; unknown | Moderate | 38 | 535 |
| SPG11 | Q96JI7 | Rare; intermediate | Definitive | 34 | 283 |
| TUBA4A | P68366 | Rare; high | Moderate | 32 | 95 |
| SIGMAR1 | Q99720 | Rare; unknown | Not available | 27 | 202 |
| CAV1 | Q03135 | Rare; small | Limited | 14 | 6065 |
| ACSL5 | Q9ULC5 | Common; small | Not available | 4 | 192 |
| LGALS1 | P09382 | Rare; high | Not available | 3 | 308 |
Table 2
Galectin-1 antibodies tested in this study.
| Company | Catalog number | Lot number | RRID (antibody registry) | Clonality | Clone ID | Host | Concentration (µg/µL) | Vendors recommended applications |
|---|---|---|---|---|---|---|---|---|
| Abcam | ab108389* | 1018000-29 | AB_10863417 | Recombinant mono | EPR3205 | Rabbit | 1.35 | WB, IF |
| Abcam | ab138513† | 1086382-8 | AB_2894851 | Monoclonal | EPR3206(2) | Rabbit | 0.26 | WB, IF |
| Abcam | ab192842* | 1011456-2 | AB_3725515 | Recombinant mono | SP247 | Rabbit | 0.04 | WB |
| ABclonal | A21218* | 3522042922 | AB_3725523 | Recombinant mono | ARC53156 | Rabbit | 1.20 | WB |
| Aviva Systems Biology | ARP58491_P050 | QC25810-42978 | AB_2046045 | Polyclonal | - | Rabbit | 0.50 | WB |
| Bio-Techne (R&D Systems) | MAB1152† | CJKC0325011 | AB_3657585 | Monoclonal | 933133 | Mouse | 0.50 | WB |
| Cell Signaling Technology | 12936* | 3 | AB_2798065 | Recombinant mono | D608T | Rabbit | 0.10 | WB, IF |
| Thermo Fisher Scientific | MA5-32779* | 79527780 | AB_2810056 | Recombinant mono | JM13-37 | Rabbit | 1.00 | WB, IP, IF |
| Thermo Fisher Scientific | MA5-35566* | 79531623 | AB_2849466 | Recombinant mono | 7W5Q6 | Rabbit | 2.00 | WB, IF |
| Thermo Fisher Scientific | MA5-47904* | 79531680 | AB_3074348 | Recombinant mono | SAIC-25B-112 | Rabbit | 1.00 | WB |
-
*
Recombinant antibody.
-
†
Monoclonal antibody.
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Antibody | HRP-Goat polyclonal anti-Rabbit antibody Secondary Antibody (H+L) | Thermo Fisher Scientific | CAT#65-6120; RRID:AB_2533967 | 0.05 µg/mL |
| Antibody | HRP-Goat polyclonal anti-Mouse antibody Secondary Antibody (H+L) | Thermo Fisher Scientific | CAT#62-6520; RRID:AB_2533947 | 0.1 µg/mL |
| Antibody | IRDye 680CW Goat polyclonal anti-Rabbit Secondary Antibody (H+L) | LI-COR Biosciences | CAT#926-68071; RRID:NA | 0.05 µg/mL |
| Antibody | IRDye 680CW Goat polyclonal anti-Mouse Secondary Antibody (H+L) | LI-COR Biosciences | CAT#926-68070; RRID:NA | 0.1 µg/mL |
| Antibody | IRDye 800CW Goat polyclonal anti-Rabbit Secondary Antibody (H+L) | LI-COR Biosciences | CAT#926-32211; RRID:NA | 0.05 µg/mL |
| Antibody | IRDye 800CW Goat polyclonal anti-Mouse Secondary Antibody (H+L) | LI-COR Biosciences | CAT#926-32210; RRID:NA | 0.1 µg/mL |
| Antibody | HRP-Goat multiclonal recombinant anti-Rabbit Secondary Antibody (H+L) | Proteintech | CAT#RGAR001; RRID:AB_3073505 | 0.05 µg/mL |
| Antibody | HRP-Goat multiclonal recombinant anti-Mouse Secondary Antibody (H+L) | Proteintech | CAT#RGAM001; RRID:AB_3068333 | 0.5 µg/mL |
| Antibody | CoraLite Plus 555-Goat multiclonal recombinant Anti-Rabbit Secondary Antibody (H+L) | Proteintech | CAT# RGAR003; RRID:AB_3073507 | 0.5 µg/mL |
| Antibody | CoraLite Plus 555-Goat multiclonal recombinant Anti-Mouse Secondary Antibody (H+L) | Proteintech | CAT# RGAM003; RRID:AB_3068539 | 0.5 µg/mL |
| Antibody | Rabbit recombinant monoclonal anti-Galectin-1 (clone EPR3205) | Abcam | CAT#ab108389; RRID:AB_10863417 | WB:1/100; IP 2 µg; IF:1/100 |
| Antibody | Rabbit recombinant monoclonal anti-Galectin-1 (clone EPR3206(2)) | Abcam | CAT#ab138513; RRID:AB_2894851 | WB:1/10,000; IP 2 µg; IF:1/250 |
| Antibody | Rabbit recombinant monoclonal anti-Galectin-1 (clone SP247) | Abcam | CAT#ab192842; RRID:AB_3725515 | WB:1/100; IP 2 µg; IF:1/100 |
| Antibody | Rabbit recombinant monoclonal anti-Galectin-1 (clone ARC53156) | ABclonal | CAT#A21218; RRID:AB_3725523 | WB:1/11,000; IP 2 µg; IF:1/500 |
| Antibody | Rabbit polyclonal anti-Galectin-1 | Aviva Systems Biology | CAT#ARP58491_P050; RRID:AB_2046045 | WB:1/100; IP 2 µg; IF:1/250 |
| Antibody | Mouse monoclonal anti-Galectin-1 (clone 933133) | Bio-Techne (R&D Systems) | CAT# MAB1152; RRID:AB_3657585 | WB:1/250; IP 2 µg; IF:1/500 |
| Antibody | Rabbit recombinant monoclonal anti-Galectin-1 (clone D608T) | Cell Signaling Technology | CAT#12936; RRID:AB_2798065 | WB:1/1000; IP 2 µg; IF:1/50 |
| Antibody | Rabbit recombinant monoclonal anti-Galectin-1 (clone JM13-37) | Thermo Fisher Scientific | CAT#MA5-32779; RRID:AB_2810056 | WB:1/2000; IP 2 µg; IF:1/2000 |
| Antibody | Rabbit recombinant monoclonal anti-Galectin-1 (clone A7W5Q6) | Thermo Fisher Scientific | CAT#MA5-35566; RRID:AB_2849466 | WB:1/200; IP 2 µg; IF:1/100 |
| Antibody | Rabbit recombinant monoclonal anti-Galectin-1 (clone SAIC-25B-112) | Thermo Fisher Scientific | CAT#MA5-47904; RRID:AB_3074348 | WB:1/1000; IP 2 µg; IF:1/1000 |
| Antibody | Rabbit recombinant monoclonal anti-CHMP2B (clone JE54-35) | Thermo Fisher Scientific | CAT#MA5-36184; RRID:AB_2890433 | WB:1/500 |
| Antibody | Rabbit recombinant monoclonal anti-FIG4 (clone N202/7R) | Addgene | CAT#192037; RRID:AB_2916241 | WB:1/100 |
| Antibody | Mouse monoclonal anti-FUS (clone CL0190) | Abcam | CAT#ab154141; RRID:AB_2885092 | WB:1/1000 |
| Antibody | Mouse monoclonal anti-HNRNPA1 (clone 4B10) | Bio-Techne (Novus Biologicals) | CAT#NB100-672; RRID:AB_524308 | WB:1/1000 |
| Antibody | Rabbit recombinant monoclonal anti-MATR3 (clone EPR10634(B)) | Abcam | CAT#ab151739; RRID:AB_2885091 | WB:1/10,000 |
| Antibody | Rabbit polyclonal anti-NEK1 | GeneTex | CAT#GTX130828; RRID:AB_2886357 | WB:1/500 |
| Antibody | Mouse monoclonal anti-OPTN (clone 6C1H4) | Proteintech | CAT#60293-1-Ig; RRID:AB_2881408 | WB:1/5000 |
| Antibody | Rabbit recombinant monoclonal anti-SIGMAR1 (clone D4J2E) | Cell Signaling Technology | CAT#61994; RRID:AB_2799617 | WB:1/1000 |
| Antibody | Mouse monoclonal anti-SOD1 (clone 2F10G1) | Proteintech | CAT#67480-1-Ig; RRID:AB_2882707 | Chemiluminescent WB:1/10,000 |
| Antibody | Rabbit polyclonal anti-SOD1 | GeneTex | CAT#GTX100554; RRID:AB_10618670 | Fluorescent WB:1/1000 |
| Antibody | Rabbit recombinant monoclonal anti-SQSTM1 (clone EPR4844) | Abcam | CAT#ab109012; RRID:AB_2810880 | Fluorescent WB:1/1000 |
| Antibody | Mouse monoclonal anti-SQSTM1 (clone 2C11) | Abcam | CAT#ab56416; RRID:AB_945626 | Chemiluminescent WB:1/1000 |
| Antibody | Rabbit recombinant monoclonal anti-TAF15 (clone JE61-92) | Thermo Fisher Scientific | CAT#MA5-44822; RRID:AB_2931279 | WB:1/1000 |
| Antibody | Rabbit recombinant monoclonal anti-TARDBP (clone 11N20) | Proteintech | CAT#80001-1-RR; RRID:AB_2882933 | WB:1/10,000 |
| Antibody | Rabbit recombinant monoclonal anti-TBK1 (clone EP611Y) | Abcam | CAT#ab40676; RRID:AB_776632 | Chemiluminescent WB:1/10,000 |
| Antibody | Rabbit recombinant monoclonal anti-TBK1 (clone E8I3G) | Cell Signaling Technology | CAT#38066; RRID:AB_2827657 | Fluorescent WB:1/1000 |
| Antibody | Rabbit recombinant monoclonal anti-TIA1 (clone D1Q3K) | Cell Signaling Technology | CAT#86050; RRID:AB_2800070 | WB:1/1000 |
| Antibody | Mouse monoclonal anti-UBQLN2 (clone 6H9) | Abcam | CAT#ab190283; RRID:AB_2747782 | WB:1/1000 |
| Antibody | Rabbit recombinant monoclonal anti-VCP (clone 7F3) | Cell Signaling Technology | CAT#2649; RRID:AB_2214629 | Chemiluminescent WB:1/10,000 |
| Antibody | Rabbit recombinant monoclonal anti-VCP (clone EPR3307(2)) | Abcam | CAT#ab109240; RRID:AB_10862588 | Fluorescent WB:1/500 |
| Antibody | Rabbit polyclonal anti-ANXA11 | Aviva Systems Biology | CAT#ARP87950_P050; RRID:AB_2894925 | WB:1/1000 |
| Antibody | Mouse monoclonal anti-C9orf72 (clone GT1553) | GeneTex | CAT#GTX634482; RRID:AB_2784545 | WB:1/1000 |
| Antibody | Rabbit polyclonal anti-CHCHD10 | Proteintech | CAT#25671-1-AP; RRID:AB_2880187 | Chemiluminescent WB:1/500 |
| Antibody | Mouse monoclonal anti-CHCHD10 (clone OTI3B8) | Thermo Fisher Scientific | CAT#MA5-27535; RRID:AB_2724132 | Fluorescent WB:1/500 |
| Antibody | Rabbit recombinant monoclonal anti-LGALS1 (clone EPR3206(2)) | Abcam | CAT#ab138513; RRID:AB_2894851 | WB:1/500 |
| Antibody | Rabbit recombinant monoclonal anti-PFN1 (clone ARC1469) | ABclonal | CAT#A9188; RRID:AB_2863682 | WB:1/10,000 |
| Antibody | Rabbit recombinant monoclonal anti-SPG11 (clone 2H25L11) | Thermo Fisher Scientific | CAT#702776; RRID:AB_2725406 | WB:1/200 |
| Antibody | Mouse monoclonal anti-ACSL5 (clone 5H8) | Abnova | CAT#H00051703-M01; RRID:AB_509241 | WB:1/200 |
| Antibody | Rabbit recombinant monoclonal anti-CAV1 (clone SZ02-01) | Thermo Fisher Scientific | CAT#MA5-32020; RRID:AB_2809314 | Chemiluminescent WB:1/500 |
| Antibody | Rabbit recombinant monoclonal anti-CAV1 (clone ARC50848) | ABclonal | CAT#A19006; RRID:AB_2862498 | Fluorescent WB:1/500 |
| Antibody | Rabbit recombinant monoclonal anti-ATXN2 (clone EPR23630-49) | Abcam | CAT#ab254362; RRID:AB_2885130 | WB:1/1000 |
| Antibody | Rabbit recombinant monoclonal anti-TUBA4A (clone EPR13478(B)) | Abcam | CAT#ab176560; RRID:AB_2860019 | Chemiluminescent WB:1/500 |
| Antibody | Rabbit recombinant monoclonal anti-TUBA4A (clone JM73-24) | Thermo Fisher Scientific | CAT#MA5-32738; RRID:AB_2810015 | Fluorescent WB:1/1000 |
| Antibody | Rabbit recombinant monoclonal anti-VAPB (clone EPR27026-50) | Abcam | CAT#ab315013; RRID:AB_3086770 | Chemiluminescent WB:1/500 |
| Antibody | Rabbit polyclonal anti-VAPB | Thermo Fisher Scientific | CAT#PA5-53023; RRID:AB_2649391 | Fluorescent WB:1/200 |
| Antibody | Rabbit recombinant monoclonal anti-ALS2 (clone EPR11185) | Abcam | CAT#ab170896; RRID:AB_2885099 | WB:1/1000 |
| Antibody | Mouse monoclonal anti-HNRNPA2B1 (clone DP3B3) | Bio-Techne (Novus Biologicals) | CAT#NB120-6102; RRID:AB_790226 | WB:1/500 |
| Antibody | Rabbit recombinant monoclonal anti-KIF5A (clone 241124H1) | Proteintech | CAT#84013-4-RR; RRID:AB_3671581 | WB:1/500 |
| Antibody | Rabbit recombinant monoclonal anti-Synaptotagmin 1 (clone HL1626) | GeneTex | CAT#GTX637119; RRID:AB_3068350 | WB:1/500 |
| Antibody | Rabbit recombinant monoclonal anti-CD44 (clone EPR18668) | Abcam | CAT#ab189524; RRID:AB_2885107 | WB:1/1000 |
| Antibody | Rabbit recombinant monoclonal anti-ISL1 (clone EPR10362) | Abcam | CAT#ab178400; RRID:AB_2927537 | WB:1/500 |
| Antibody | Rat recombinant monoclonal anti-Myelin Basic Protein antibody (clone 12) | Abcam | CAT#ab7349; RRID:AB_305869 | WB:1/500 |
| Antibody | Rabbit polyclonal anti-Tyrosine Hydroxylase | Pel-Freeze | CAT#P40101-150: RRID:AB_2617184 | WB:1/500 |
| Antibody | Rabbit recombinant monoclonal anti-CD11b (clone EPR1344) | Abcam | CAT#ab133357; RRID:AB_2650514 | WB:1/500 |
| Antibody | Rabbit recombinant monoclonal anti-IBA1 (clone E4O4W) | Cell Signaling Technology | CAT#17198; RRID:AB_2820254 | WB:1/500 |
| Antibody | Chicken monoclonal recombinant anti-IBA1 (clone Ch311H9) | Synaptic Systems | CAT#234009; RRID:AB_2891282 | IF:1/1000 |
| Antibody | Mouse monoclonal anti-CD11b, pre-conjugated with PE (clone ICRF44) | BD Biosciences | CAT#555388; RRID:AB_395789 | IF:1/20 |
| Antibody | Mouse monoclonal anti-CD45, pre-conjugated with Alexa Fluor 700 (clone HI30) | BioLegend | CAT#304024; RRID:AB_493761 | IF:1/40 |
| Antibody | Rabbit polyclonal anti-PU.1 | Cell Signaling Technology | CAT#2266; RRID:AB_10692379 | IF:1/500 |
| Commercial assay or kit | RNeasy Mini Kit | QIAGEN | CAT#74106 | |
| Commercial assay or kit | STEMdiff Forebrain Neuron Differentiation Kit | STEMCELL Technologies | CAT#08600 | |
| Commercial assay or kit | STEMdiff Forebrain Neuron Maturation Kit | STEMCELL Technologies | CAT#08605 | |
| Commercial assay or kit | STEMdiff Hematopoietic Kit | STEMCELL Technologies | CAT#05310 | |
| Commercial assay or kit | STEMdiff Hematopoietic Kit | STEMCELL Technologies | CAT#05310 | |
| Commercial assay or kit | STEMdiff Hematopoietic Kit | STEMCELL Technologies | CAT#05310 | |
| Cell line (human) | HAP1 WT | Horizon Discovery | CAT#C631; RRID:CVCL_Y019 | Cells were confirmed mycoplasma-free |
| Cell line (human) | HAP1 LGALS1 KO | Horizon Discovery | CAT#HZGHC005989c006; RRID:CVCL_XQ14 | Cells were confirmed mycoplasma-free |
| Cell line (human) | Healthy control iPSCs from the AIW002-02 background | EDDU, McGill University | NA | Cells were confirmed mycoplasma-free |
| Cell line (human) | Healthy control iPSCs from the QPN929 background | EDDU, McGill University | NA | Cells were confirmed mycoplasma-free |
| Software, algorithm | LI-COR Image Studio Lite | licorbio.com/image-studio | NA | |
| Software, algorithm | FIJI Software | fiji.sc/ | RRID:SCR_002285 | |
| Software, algorithm | MetaXpress High-Content Screening Software version 6.7.1.157 | moleculardevices.com/products/cellular-imaging-systems/high-content-analysis/metaxpress - overview | RRID:SCR_016654 | |
| Software, algorithm | PyCharm 2023.3.4 | jetbrains.com/pycharm/ | RRID:SCR_018221 | |
| Software, algorithm | RStudio 2023.09.1 | cran.rstudio.com/ | RRID:SCR_000432 | |
| Other | Accutase | STEMCELL Technologies | CAT#07922 | |
| Other | Antibody Registry | antibodyregistry.org | RRID:SCR_006397 | |
| Other | Astrocyte Growth Supplements (AGS) | ScienCell Research Laboratories | CAT#1852 | |
| Other | Astrocyte Medium (AM) | ScienCell Research Laboratories | CAT#1801-b | |
| Other | BLUelf Prestained Protein Ladder | FroggaBio | CAT#PM008 | |
| Other | Cellosaurus—- a knowledge resource on cell lines | cellosaurus.org | RRID:SCR_013869 | |
| Other | CellTracker deep red dye | Thermo Fisher Scientific | CAT#C34565 | |
| Other | CellTracker green chloromethyl derivative of fluorescein diacetate (CMFDA) dye | Thermo Fisher Scientific | CAT#C2925 | |
| Other | Centrifugal filter unit, Amicon Ultra-15 | MilliporeSigma | CAT#UFC901096 | |
| Other | CiteAb—Reagent Search & Life Science Data | citeab.com | RRID:SCR_009653 | |
| Other | ClinGen | https://clinicalgenome.org/ | RRID:SCR_014968 | |
| Other | DepMap: The Cancer Dependency Map Project at Broad Institute | depmap.org/portal | RRID:SCR_017655 | |
| Other | Live/Dead Fixable Aqua viability dye | Thermo Fisher Scientific | CAT#L34965 | |
| Other | mTeSR1 | STEMCELL Technologies | CAT#85850 | |
| Other | NuPAGE Bis-Tris Midi Protein Gels, 10%, 1.0 mm | Thermo Fisher Scientific | CAT#WG1201BOX | |
| Other | NuPAGE MES SDS Running Buffer (20×) | Thermo Fisher Scientific | CAT#NP000202 | |
| Other | NuPAGE MOPS SDS Running Buffer (20×) | Thermo Fisher Scientific | CAT#NP000102 | |
| Other | PhenoPlate, 96-well, optically clear flat bottom | PerkinElmer | CAT#6055300 | |
| Other | Pierce IP Lysis Buffer | Thermo Fisher Scientific | CAT#87788 | |
| Other | PubMed | pubmed.ncbi.nlm.nih.gov | RRID:SCR_004846 | |
| Other | RIPA Lysis and Extraction Buffer | Thermo Fisher Scientific | CAT#89901 | |
| Other | Structural Genomics Consortium | https://www.thesgc.org/ | RRID:SCR_003890 | |
| Other | The Human Protein Atlas | proteinatlas.org | RRID:SCR_006710 | |
| Other | TRIzol | Thermo Fisher Scientific | CAT#15596026 | |
| Other | TrueStain blocking reagent | BioLegend | CAT#422302 | |
| Other | Zenodo | https://zenodo.org/ | RRID:SCR_004129 |
Additional files
-
Supplementary file 1
List of 303 antibodies targeting ALS-RAP proteins evaluated in this study, together with the DOI linking to the corresponding antibody characterization reports.
- https://cdn.elifesciences.org/articles/111349/elife-111349-supp1-v1.xlsx
-
Supplementary file 2
Sequences of synthetic ScFv antibodies.
- https://cdn.elifesciences.org/articles/111349/elife-111349-supp2-v1.xlsx
-
MDAR checklist
- https://cdn.elifesciences.org/articles/111349/elife-111349-mdarchecklist1-v1.pdf
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
A validated antibody toolbox for ALS research
eLife 15:RP111349.
https://doi.org/10.7554/eLife.111349.3
