Fixation is a process that helps to lock proteins in place on cells you plan to analyze. There can be a variety of reasons to fix your cells, including:

  • Timing/Covenience (experiments that cannot be completed in one day)
  • Safety (biohazardous samples that must be fixed/killed prior to analysis)
  • Intracellular analysis of targets (also includes permeabilization)

Fixation is a common practice, but it creates some difficulties if samples are fixed prior to staining for surface antibodies. Fixation can alter the epitope, making it difficult for your antibody to recognize its target. The information provided below is based on our in-house testing of our clones on unfixed cells (no fixation) and cells fixed with 4% paraformaldehyde prior to staining (post-fixation). We have provided data where possible. Black histograms/dot plots represent unstained samples. Purple histograms/dot plots represent staining with the indicated test antibody.

 

Please note that this data is based on several variables, including the types of cells analyzed, the antibody's fluorophore, and relative abundance of the antigen. Costains are also recommended to help you identify your population of interest. If you are planning to use a live/dead stain (such as Zombie Dyes), stain with them prior to fixation.

 

While these results will not necessarily guarantee the success or failure of staining a fixed epitope, it will at least provide you with an idea of how your samples might look.

 

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Specificity Clone Compatability Sample Costain Data Products
CCR8 L263G8 No Lysed whole blood, lymphocytes   View Data View Products
CCR10 6588-5 No Lysed whole blood, lymphocytes   View Data View Products
CD3 UCHT1 Yes Lysed whole blood, lymphocytes   View Data View Products
CD3 OKT3 Yes Lysed whole blood     View Products
CD3 HIT3a Yes Lysed whole blood     View Products
CD3 SK7 Yes Lysed whole blood     View Products
CD4 OKT4 Yes Lysed whole blood     View Products
CD4 RPA-T4 Yes Lysed whole blood     View Products
CD5 UCHT2 Yes Lysed whole blood     View Products
CD6 BL-CD6 Yes Lysed whole blood, lymphocytes   View Data View Products
CD7 CD7-6B7 Yes Lysed whole blood, lymphocytes   View Data View Products
CD8 RPA-T8 Yes Lysed whole blood     View Products
CD8 HIT8a Yes Lysed whole blood     View Products
CD11a HI111 Yes Lysed whole blood, lymphocytes   View Data View Products
CD11b ICRF44 Yes Lysed whole blood     View Products
CD11c 3.9 No Lysed whole blood, monocytes   View Data View Products
CD13 WM15 Yes Lysed whole blood     View Products
CD14 M5E2 Yes Lysed whole blood, monocytes   View Data View Products
CD14 HCD14 Yes Lysed whole blood     View Products
CD15 W6D3 Yes Lysed whole blood     View Products
CD15 HI98 Yes Lysed whole blood, granulocytes   View Data View Products
CD16 3G8 Yes Lysed whole blood     View Products
CD18 TS1/18 Yes Lysed whole blood     View Products
CD19 HIB19 Yes Lysed whole blood, lymphocytes   View Data View Products
CD23 EBVCS-5 Yes Lysed whole blood     View Products
CD24 ML5 No Lysed whole blood, lymphocytes   View Data View Products
CD25 BC96 No Lysed whole blood     View Products
CD27 O323 Yes Lysed whole blood     View Products
CD27 M-T271 Yes Lysed whole blood, lymphocytes   View Data View Products
CD28 CD28.2 Yes Lysed whole blood     View Products
CD31 WM59 Yes Lysed whole blood     View Products
CD34 581 No Lysed whole blood     View Products
CD38 HIT2 Yes Lysed whole blood     View Products
CD40 5C3 Yes Lysed whole blood     View Products
CD45 HI30 Yes Lysed whole blood     View Products
CD47 CC2C6 Yes Lysed whole blood, lymphocytes   View Data View Products
CD50 CBR-IC3/1 Yes Lysed whole blood     View Products
CD56 MEM-188 Yes Lysed whole blood, lymphocytes   View Data View Products
CD56 HCD56 Yes Lysed whole blood, lymphocytes   View Data View Products
CD62L DREG-56 Yes Lysed whole blood, lymphocytes   View Data View Products
CD86 IT2.2 Yes Lysed whole blood, monocytes   View Data View Products
CD88 S5/1 Yes Lysed whole blood, granulocytes   View Data View Products
CD95 DX2 Yes Lysed whole blood, lymphocytes   View Data View Products
CD99 HCD99 Yes Lysed whole blood, lymphocytes   View Data View Products
CD127 (IL-7Rα) A019D5 No Lysed whole blood, lymphocytes   View Data View Products
CD134 Ber-ACT35 (ACT35) No Lysed whole blood     View Products
CD172a/b SE5A5 Yes Lysed whole blood     View Products
CD172g LSB2.20 Yes Lysed whole blood     View Products
CD181 (CXCR1) 8F1/CXCR1 No Lysed whole blood, granulocytes. Lymphocytes and monocytes display high non-specific binding.   View Data View Products
CD182 (CXCR2) 5E8/CXCR2 No Lysed whole blood, granulocytes   View Data View Products
CD183 (CXCR3) G025H7 No Lysed whole blood, lymphocytes   View Data View Products
CD184 (CXCR4) 12G5 Yes Lysed whole blood, lymphocytes   View Data View Products
CD193 (CCR3) 5E8 No Lysed whole blood, granulocytes   View Data View Products
CD194 (CCR4) L291H4 No Lysed whole blood, lymphocytes   View Data View Products
CD195 (CCR5) T21/8 No Lysed whole blood, lymphocytes   View Data  
CD195 (CCR5) HEK/1/85a No Lysed whole blood, lymphocytes   View Data View Products
CD195 (CCR5) J418F1 No Lysed whole blood, lymphocytes   View Data View Products
CD196 (CCR6) G034E3 No Lysed whole blood, lymphocytes   View Data View Products
CD197 (CCR7) G043H7 No Lysed whole blood, lymphocytes   View Data View Products
CD199 (CCR9) L053E8 No Lysed whole blood, lymphocytes   View Data View Products
CD244 (2B4) C1.7 Yes Lysed whole blood     View Products
CD268 (BAFF-R) 11C1 Yes Lysed whole blood     View Products
CD279 EH12.2H7 No Lysed whole blood, lymphocytes   View Data View Products
CD282 (TLR2) TL2.1 Yes Lysed whole blood     View Products
CX3CR1 2A9-1 No Lysed whole blood, lymphocytes   View Data View Products
CX3CR1 K0124E1 No Lysed whole blood, lymphocytes   View Data View Products
HLA-DR L243 Yes Lysed whole blood     View Products
IgM MHM-88 No Lysed whole blood     View Products

CCR8 (L263G8)

CCR10 (6588-5)

CD3 (UCHT1)

CD6 (BL-CD6)

CD7 (CD7-6B7)

CD11a (HI111)

CD11c (3.9)

CD14 (M5E2)

CD15 (HI98)

CD19 (HIB19)

CD24 (ML5)

CD27 (M-T271)

CD47 (CC2C6)

CD56 (HCD56)

CD56 (MEM-188)

CD62L (DREG-56)

CD86 (IT2.2)

CD88 (S5/1)

CD95 (DX2)

CD99 (HCD99)

CD127 (IL-7Rα) (A019D5)

CD181 (CXCR1) (8F1/CXCR1)

CD182 (CXCR2) (5E8/CXCR2)

CD183 (CXCR3) (G025H7)

CD184 (CXCR4) (12G5)

CD193 (CCR3) (5E8)

CD194 (CCR4) (L291H4)

CD195 (CCR5) (HEK/1/85a)

CD195 (CCR5) (J418F1)

CD195 (CCR5) (T21/8)

CD196 (CCR6) (G034E3)

CD197 (CCR7) (G043H7)

CD199 (CCR9) (L053E8)

CD279 (EH12.2H7)

CX3CR1 (2A9-1)

CX3CR1 (K0124E1)

Specificity Clone Compatability Sample Costain Data Products
CD1d 1B1 No C57BL/6 Spleen     View Products
CD3 17A2 Yes C57BL/6 Spleen   View Data View Products
CD3 145-2C11 Yes C57BL/6 Spleen   View Data View Products
CD4 RM4-5 Yes C57BL/6 Spleen   View Data View Products
CD4 GK1.5 Yes C57BL/6 Spleen   View Data View Products
CD4 RM4-4 No C57BL/6 Spleen   View Data View Products
CD5 53-7.3 Yes C57BL/6 Spleen   View Data View Products
CD8a 53-6.7 Yes C57BL/6 Spleen   View Data View Products
CD8b YTS156.7.7 Yes C57BL/6 Spleen   View Data View Products
CD8b.2 53-5.8 No C57BL/6 Spleen   View Data View Products
CD11b M1/70 Yes C57BL/6 Bone Marrow   View Data View Products
CD11c N418 No C57BL/6 Spleen     View Products
CD16/32 93 No C57BL/6 Spleen     View Products
CD19 6D5 Yes C57BL/6 Spleen   View Data View Products
CD21/35 7E9 Yes C57BL/6 Spleen   View Data View Products
CD23 B3B4 No C57BL/6 Spleen   View Data View Products
CD24 M1/69 No C57BL/6 Spleen     View Products
CD27 LG.3A10 Yes C57BL/6 Spleen   View Data View Products
CD28 37.51 No C57BL/6 Spleen     View Products
CD29 HMβ1-1 Yes C57BL/6 Spleen   View Data View Products
CD31 MEC13.3 Yes C57BL/6 Spleen   View Data View Products
CD31 390 No C57BL/6 Spleen   View Data View Products
CD40 HM40-3 No C57BL/6 Spleen   View Data View Products
CD40 3/23 No C57BL/6 Spleen   View Data View Products
CD44 IM7 Yes C57BL/6 Spleen   View Data View Products
CD45 30-F11 Yes C57BL/6 Spleen   View Data View Products
CD45.2 104 Yes C57BL/6 Spleen   View Data View Products
CD45R (B220) RA3-6B2 Yes C57BL/6 Spleen   View Data View Products
CD48 HM48-1 Yes C57BL/6 Spleen   View Data View Products
CD49b HMα2 No C57BL/6 Spleen     View Products
CD62L MEL-14 No C57BL/6 Spleen   View Data View Products
CD71 RI7217 No C57BL/6 Bone Marrow   View Data View Products
CD81 EAT-2 No C57BL/6 Spleen     View Products
CD88 (C5aR) 20/70 Yes C57BL/6 Bone Marrow   View Data View Products
CD103 2E7 No C57BL/6 Spleen     View Products
CD117 (c-kit) 2B8 No C57BL/6 Bone Marrow     View Products
CD122 TM-β1 Yes C57BL/6 Spleen   View Data View Products
CD127 (IL-7Rα) A7R34 No C57BL/6 Spleen   View Data View Products
CD135 A2F10 No C57BL/6 Bone Marrow     View Products
CD138 281-2 Yes C57BL/6 Spleen     View Products
CD150 TC15-12F12.2 No C57BL/6 Spleen   View Data View Products
CD183 (CXCR3) CXCR3-173 Yes C57BL/6 Spleen   View Data View Products
CD196 (CCR6) 29-2L17 No C57BL/6 Spleen     View Products
CD197 (CCR7) 4B12 No C57BL/6 Spleen     View Products
CD205 (DEC-205) NLDC-145 No C57BL/6 Bone Marrow     View Products
CD210 (IL-10R) 1B1.3a No C57BL/6 Spleen     View Products
CD274 10F.9G2 Yes C57BL/6 Spleen   View Data View Products
CD317 927 Yes C57BL/6 Spleen     View Products
CD335 29A1.4 Yes C57BL/6 Spleen     View Products
Gr1 RB6-8C5 Yes C57BL/6 Bone Marrow   View Data View Products
H-2kb AF6-88.5 Yes C57BL/6 Spleen   View Data View Products
I-A/I-E M5/114.15.2 Yes C57BL/6 Spleen   View Data View Products
I-Ab AF6-120.1 No C57BL/6 Spleen   View Data View Products
IgD 11-26c.2a Yes C57BL/6 Spleen   View Data View Products
IgM RMM-1 Yes C57BL/6 Spleen PE: Clone RMM-1, Anti-mouse IgM; APC: Clone 6D5, Anti-mouse CD19 View Data View Products
KLRG1 (MAFA) 2F1/KLRG1 Yes C57BL/6 Spleen     View Products
LPAM-1 (Integrin α4β7) DATK32 No C57BL/6 Spleen     View Products
Ly6C HK1.4 Yes C57BL/6 Bone Marrow   View Data View Products
Ly6G 1A8 Yes C57BL/6 Bone Marrow   View Data View Products
NK1.1 PK136 Yes C57BL/6 Spleen PE: Clone PK136, Anti-mouse NK1.1; APC: Clone DX5, Anti-mouse CD49b View Data View Products
Sca-1 (Ly6A/E) E13-161.7 Yes C57BL/6 Spleen   View Data View Products
Sca-1 (Ly6A/E) D7 Yes C57BL/6 Spleen   View Data View Products
Siglec H 551 No C57BL/6 Spleen     View Products
TCRβ H57-597 Yes C57BL/6 Spleen   View Data View Products
TER-119 TER-119 Yes C57BL/6 Bone Marrow   View Data View Products

CD3 (145-2C11)

CD3 (17A2)

CD4 (GK1.5)

CD4 (RM4-4)

CD4 (RM4-5)

CD5 (53-7.3)

CD8a (53-6.7)

CD8b (YTS156.7.7)

CD8b.2 (53-5.8)

CD11b (M1/70)

CD19 (6D5)

CD21/35 (7E9)

CD23 (B3B4)

CD27 (LG.3A10)

CD29 (HMβ1-1)

CD31 (390)

CD31 (MEC13.3)

CD40 (3/23)

CD40 (HM40-3)

CD44 (IM7)

CD45 (30-F11)

CD45.2 (104)

CD45R (B220) (RA3-6B2)

CD48 (HM48-1)

CD62L (MEL-14)

CD71 (RI7217)

CD88 (C5aR) (20/70)

CD122 (TM-β1)

CD127 (IL-7Rα) (A7R34)

CD150 (TC15-12F12.2)

CD183 (CXCR3) (CXCR3-173)

CD274 (10F.9G2)

Gr1 (RB6-8C5)

H-2kb(AF6-88.5)

I-A/I-E (M5/114.15.2)

I-Ab(AF6-120.1)

IgD (11-26c.2a)

IgM (RMM-1)

Ly6C (HK1.4)

Ly6G (1A8)

NK1.1 (PK136)

Sca-1 (Ly6A/E) (D7)

Sca-1 (Ly6A/E) (E13-161.7)

TCRβ (H57-597)

TER-119 (TER-119)

Chemokine receptors often display high false positives across lymphocytes, monocytes, and granulocytes when staining post-fixation.

 

Lymphocyte, monocyte, and granulocyte gates are based on FSC and SSC profiles.

 

FL-1 indicates the FITC Channel


Learn more about our fixation solutions:


True-Phos™ Perm Buffer is specially designed for the flow cytometric staining of phospho-specific targets like pSTAT3. 

Fixation Buffer – for use with Permeabilization Wash Buffer (10X) to detect intracellular targets such as cytokines, chemokines, or other cytoplasmic antigens.

FluoroFix™ Buffer – for fixation of immunofluorescence stained cells, optimized to stabilize tandem dyes.

True-Nuclear™ Transcription Factor Buffer Set - for optimal staining of intranuclear and cytoplasmic antigens. Cytokine staining may not be optimal with this buffer as loss from cells will be evident with this method of fixation and permeabilization.  Refer to our Fixation and Permeabilization buffers above for cytokine staining.

RBC Lysis/Fixation Solution (10X) - optimized for one-step lysing of red blood cells (RBCs) and fixation of the remaining blood cells following immunofluorescent staining with fluorochrome-conjugated antibodies, suitable for either lyse/wash or lyse/no wash staining procedures.

 

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